Water Dynamics and Its Role in Structural Hysteresis of Dissolved

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Water dynamics and its role in structural hysteresis of dissolved organic matter Pellegrino Conte, and Jiri Ku#erík Environ. Sci. Technol., Just Accepted Manuscript • DOI: 10.1021/acs.est.5b04639 • Publication Date (Web): 26 Jan 2016 Downloaded from http://pubs.acs.org on January 29, 2016

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Environmental Science & Technology

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Water dynamics and its role in structural hysteresis of dissolved organic

2

matter

3 4

Pellegrino Conte1 and Jiri Kucerik2*

5

1

6

Scienze edificio 4, 90128, Palermo, Italy

7

2

8

Fortstrasse 7, 768 29, Landau, Germany

Dipartimento di Scienze Agrarie e Forestali, Università degli Studi di Palermo, v.le delle

Institute for Environmental Sciences, University of Koblenz-Landau, Campus Landau,

9 10 11 12 13 14 15 16 17

* Corresponding author: Jiri Kucerik, e-mail [email protected], tel. +49 6341 280 31582

18

fax +49 6341 280 31576

1 ACS Paragon Plus Environment


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Abstract

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Knowledge of structural dynamics of dissolved organic matter (DOM) is of paramount importance

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for understanding DOM stability and role in the fate of solubilized organic and inorganic

22

compounds (e.g. nutrients and pollutants) either in soils or aquatic systems. In this study, Fast

23

Field Cycling (FFC) 1H NMR relaxometry was applied to elucidate structural dynamics of terrestrial

24

DOM, represented by two structurally contrasting DOM models such as the Suwanee river (SRFA)

25

and the Pahokee peat (PPFA) fulvic acids purchased by the International Humic Substance Society.

26

Measurement of NMR relaxation rate of water protons in heating-cooling cycles revealed

27

structural hysteresis in both fulvic acids. In particular, structural hysteresis was related to the

28

delay in re-establishing water network around fulvic molecules as a result of temperature

29

fluctuations. The experiments revealed that the structural temperature dependency and

30

hysteresis were more pronounced in SRFA than in PPFA. This was attributed to the larger content

31

of hydrogel-like structure in SRFA stabilized, at a larger extent, by H-bonds between carboxylic

32

and phenolic groups. Moreover, results supported the view that terrestrial DOM consist of a

33

hydrophobic rigid core surrounded by progressively assembling amphiphilic and polar molecules,

34

which form an elastic structure that can mediate reactivity of the whole DOM.

35 36 37 38

Key words: dissolved organic matter, water hysteresis, Fast Filed Cycling NMR, correlation time,

39

relaxation, structural hysteresis, temperature variation


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40

Introduction

41

Dissolved organic matter (DOM) represents an important part of natural organic matter (NOM).

42

In terrestrial eco-systems, DOM mediates a myriad of biogeochemical mechanisms and responses

43

to changes in ecological processes.1 In particular, terrestrial DOM is involved in the solubilization

44

and transport of both inorganic2,3,4 and organic molecules and colloids,3, 2, 9

10

rock weathering, pedogenesis of topsoils and subsoils,

6, 11

5-8

micronutrient

soil water repellency12

45

availability,

46

and soil texture.13

47

Up to now, conformational arrangement of terrestrial DOM has been described by many, often

48

contradicting, models including the linear macromolecular polyelectrolyte hypothesis,14, 15 the

49

supramolecular assemblies of molecules stabilized by weak interactions,16, 17 the heterogeneous

50

Donnan gel phases,18 or, the mixture of supra and macromolecules.19 Although there is still not a

51

general consensus about DOM primary composition, DOM dynamics is attributed to its structural

52

complexity and flexibility.20, 21

53

To explain DOM ability to enhance solubility of hydrophic organic compounds (HOC),22 or its

54

capacity to decrease water surface tension,20 DOM has been assumed to form micelles akin to

55

classical surfactants.20, 23-25 The formation of micelle, i.e. the critical micelle concentration (CMC),

56

is reported in the range from 5 to 10 g L-1.26 However, in some cases, HOC solubilization was also

57

observed at concentrations far below the aforementioned CMC values.17 As an example, Kopinke

58

et al.,8 showed that for very hydrophobic compounds, dissolved humic acids show a significant

59

sorption effect at the concentration of 10 mg L-1. In addition, DOM ability to solubilize either

60

1,1,1-trichloro-2,2-bis(p-chlorophenyl) ethane or DDT appeared to decrease with increasing DOM

61

concentration.27 This anomaly cannot be explained by a classical micellar concept, but it can be

62

explained by the formation of premicellar aggregates, i.e. assemblies containing partially

63

hydrophobic regions formed below the critical micelle concentration.28-31 In addition, Drastik et

64

al.32 hypothesized that enhanced HOC solubility at low DOM concentration can be caused by a co-

65

solute effect (i.e. hydrotrophy) of the dissolved polar amphiphilic and hydrophilic molecules.

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Conte and Berns33 showed that humic and fulvic acids, considered as representatives of

67

NOM, consist of condensed aromatic hydrophobic cores surrounded by more hydrophilic

68

molecules. Recent ultrasonic velocimetry study of DOM supported this view and showed

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that hydration of such a hydrophilic envelope is concentration dependent.32 At low

70

concentration, small hydrophobic assemblies are stabilized by dissolved hydrophilic and

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amphiphilic molecules. As concentration increases, these molecules progressively form a 3 ACS Paragon Plus Environment


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physical hydrogel. Around concentration of 1 g L-1, an abrupt change in hydration

73

characteristics appears. This can be explained as a change in domination hydration

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mechanisms.32, 34 The same concentration range was observed also by analyzing adiabatic

75

compressibility of humic acids sols.35

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Similar model was also suggested by Baigorri et al.,19 who concluded that humic substances

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in solution consist of a macromolecular fraction with stable aggregates and supramolecular

78

associations of low molecular weight. Chilom and Rice26 demonstrated that the least

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abundant fraction of humic acids consisting of amphiphilic molecules facilitates aggregation

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of other aromatic and more abundant fractions. The authors also showed that only

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approximately one-third of their components displayed amphiphilic characteristics and

82

supported the idea of Guetzloff and Rice20 that not all molecules participate in micelle

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formation. Lam et al.,36 showed that carboxyl-rich alicyclic molecules play a more prominent

84

role in DOM aggregation than material derived from linear terpenoids, lignin and

85

carbohydrates.

86

Apart from concentration, pH and salinity,37 DOM structure and reactivity are also affected

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by temperature 3,36. Upon heating and cooling, Jia et al.38 observed a hysteresis in sorption

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and desorption of perfluorooctane sulfonate (PFOS) in dissolved humic substances. Palmer

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and von Wandruszka39 studied the influence of temperature on the size of DOM aggregates.

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They observed an increase in DOM aggregate sizes akin to surfactant clouding at around

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40°C. On the contrary, another study32 reported two strong transitions at around 20 and

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40°C in DOM systems up to concentration of around 4 g L-1. The transitions were attributed

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to the disruption of H-bonds stabilizing the hydrogel formed by polar and amphiphilic DOM

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moieties. At all investigated concentrations (from 0.05 up to 10 g L-1), DOM exhibited

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structural hysteresis as temperature was varied, which was explained as a difference

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between the rate of structural collapse and H-bond regeneration.32 In past, hysteresis in

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ultrasonic velocities has already been observed also for pure water and its mixtures with

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other liquids.40 This implies that DOM structural hysteresis might be caused primarily by

99

water cluster dynamics. However, this hypothesis requires additional experimental

100

verifications.

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It is worth noting that the self-assembling of molecules in aqueous systems is primarily due

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to the dynamics and arrangement of surrounding water-cluster-network (i.e. water

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structure).41, 42 Therefore, it is beneficial to assess the dynamics of dissolved and aggregated 4 ACS Paragon Plus Environment

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molecules by studying the dynamics of surrounding water molecules (e.g. refs43, 44). Indeed,

105

the role of water dynamics is frequently studied for biomolecules such as proteins and DNA,

106

45

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assembling are still scarce.

108

In order to verify the view reported by Drastik et al.32 we tested the hypotheses that (i) DOM

109

with larger content of polar and amphiphilic molecules is more vulnerable to structural

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transitions and (ii) water dynamics plays an important role in DOM temperature-induced

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hysteresis. For this reason, we analyzed water behavior in two water soluble fulvic acids

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from the International Humic Substances Society (IHSS) by high performance size exclusion

113

chromatography (HPSEC) and fast field cycling (FFC) NMR relaxometry.47-51 Results showed

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that DOM structural dynamics is largely governed by H-bonds, which stabilize the hydrogel

115

formed by polar and amphiphilic molecules surrounding a hydrophobic core which is, in turn,

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made predominantly by aromatic molecules held together by van der Waals forces. The

117

experimental data also suggest that temperature induced hysteresis in DOM structure is

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caused primarily by hysteresis in water clustering, which, then, feedbacks in H-bonds

119

rearrangement delay.

synthetic polymers,43 or cations and anions,46 but relevant studies on its role in DOM self-

120 121

Materials and methods

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Samples

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Suwanee River (SRFA) and Pahokee peat (PPFA) fulvic acids (FA) were purchased from the

124

International Humic Substances Society (IHSS) and analyzed as is. Table 1 lists the selected

125

results

126

(http://www.humicsubstances.org) and in Thorn et al.52

of

the

elemental

and

structural

analysis

reported

by

the

provider

127 128

High performance size exclusion chromatography (HPSEC)

129

HPSEC experiments were performed by using a Ultimate 3000 Standard Chromatography

130

Station (Dionex Inc., USA), equipped with a 600 × 7.8 mm BioSep S2000 column

131

thermostated at 25°C, and a BioSep Guard pre-column with a 0.2-µm stainless steel inlet

132

filter (Phenomenex Inc., USA). The UV detector set at 280 nm was used for the detection.

133

The eluent was a 50 mmol L–1 water (MilliQ) solution of NaH2PO4.H2O adjusted to pH 7 by

134

using 1 mol L-1 NaOH solution. The flow rate was set at 0.5 mL min–1, the concentration of

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SRFA and PPFA solutions used for the analysis were 0.6 mg mL–1, injected volume was 100 5 ACS Paragon Plus Environment


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µL. The calibration of the column was carried out using poly(styrenesulphonates) standards

137

of 194.2, 145, 32.9, 14.9, 6.53 and 0.91 kDa mass (Polymer Standards Service Ltd., Germany).

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The weight (MW) and number (MN) averaged molecular weights and polydispersity index

139

(PDI= MW/MN) were calculated according to the ref.53 based on the semi-logarithmic

140

calibration curve of retention time and molecular weight of eluted standards. According to

141

Sutton and Sposito,16 the determined values can be considered only as apparent values, but

142

their comparison provides a valuable insight into aggregation of FAs.

143 144

Fast field cycling nuclear magnetic resonance (FFC NMR)

145

1

146

Larmor frequencies) were acquired on a Stelar Spinmaster FFC2000 Relaxometer (Stelars.r.l.;

147

Mede, PV, Italy) at several temperatures from 25 to 52°C. The details of the technique for

148

environmental applications are reported in ref.51 Briefly, proton spins were polarized at a

149

polarization field (BPOL) corresponding to a proton Larmor frequency (ωL) of 24 MHz for a

150

period of polarization (TPOL) corresponding to about five times the T1 estimated at this

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frequency. After each BPOL application, the magnetic field intensity (abbreviated as BRLX) was

152

systematically changed in the proton Larmor frequency (ωL) comprised in the range 0.01–

153

30.0 MHz. The period τ, during which BRLX was applied, has been varied on 20 logarithmic

154

spaced time sets, each of them adjusted at every relaxation field to optimize the sampling of

155

the decay/recovery curves. Free induction decays (FID) were recorded following a single 1H

156

90° pulse applied at an acquisition field (BACQ) corresponding to the proton Larmor frequency

157

of 16 MHz. A time domain of 100 µs sampled with 512 points was applied. Field switching

158

time was 3 ms, whereas spectrometer dead time was 15 µs. For all experiments a recycle

159

delay of 5 times T1 was used.

160

Both FA were prepared by their dissolution in Milli-Q grade water and homogenization using

161

a Vortex Mixer Cole Palmer for 1 minute in order to homogenize the system and to remove

162

paramagnetic oxygen, which may affect the relaxometry experiments. The concentration of

163

both solutions was chosen 40 g L-1 in order to suppress as much as possible the

164

conformation changes in FA and thus measure only changes induced by water properties.32

165

Furthermore, this concertation is far above the critical micelle concentration reported by

166

other authors e.g.20 thus, the molecules were self-assembled in thermodynamically stable

167

aggregates. The pH of both sols were around 4. The obtained sols were measured under

H nuclear magnetic resonance dispersion profiles (i.e. relaxation rates R1 or 1/T1 vs. proton


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isothermal conditions in hermetically sealed cuvettes. The measurement was started exactly

169

after 30 minutes after reaching the requested temperature by the NMR system. This

170

protocol was applied for both heating and cooling cycles. Furthermore, the results in

171

previous work showed that the changes in FA are reversible up to 90°C,32 therefore, only

172

physical structure of FA was affected under conditions used in this work.

173 174

FFC NMR data processing

175

The NMRD profiles were fitted in Origin Pro 7.5 SR6 with a Lorentzian function of the

176

type49,51:

Aiτ C ,i

n

R1 = ∑

177

1

1 + (ω Lτ C ,i )

2

(1)

178

This equation is needed in order to account for the stretched shape of the dispersion

179

curves.51 In equation (1), R1 is the longitudinal relaxation rate, ωL is the proton Larmor

180

frequency, Ai is a constant containing the proton quantum-spin number, the proton

181

magnetogyric ratio, the Planck constant, and the electron-nuclear hyperfine coupling

182

constant describing interactions between resonant protons and unpaired electrons. As a

183

matter of fact, the larger this constant, the faster is the longitudinal relaxation rate due to

184

stronger electron-nucleus interactions. τ C ,i is the correlation time of the i-th relaxing

185

component measuring the time needed for molecular re-orientation. It is a typical

186

parameter for spectral density which, in turn, describes random molecular motions.49 The

187

number n of Lorentzians that can be included in eq. (1) without unreasonably increasing the

188

number of parameters were determined by means of the Merit function analysis.54 For the

189

present paper, n=4 was used for the mathematical fit of the NMRD profiles. It must be

190

stated that the fitting parameters achieved in equation (1) do not have physical

191

significance.50 They can only be used to provide a weight average correlation time according

192

to equation (2):51 n

∑ Aτ

i i

193

τC =

1

(2)

n

∑A

i

1

194

The activation energy and frequency factor of the proton relaxation was calculated

195

according to the equation derived from the Arrhenius equation55 7 ACS Paragon Plus Environment


R1 = A exp(−

196

EA ) RT

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(3)

197

Where R1 is the proton relaxation rate, A is the frequency factor, Ea is the activation energy,

198

R is the gas constant, and T is the thermodynamic temperature.

199 200

Results and Discussion

201

HPSEC evaluation of PPFA and SRFA characteristics

202

Figure 1 reports the HPSEC profiles of PPFA and SRFA. To illustrate the distribution of

203

molecular weights, the retention time was recalculated into molecular weight scale. In

204

particular, PPFA revealed a more intense band in the high molecular weight range (∼ 9 kg

205

mol-1) and weaker bands in the low molecular weight interval (∼4 kg mol-1 and ∼2 kg mol-1)

206

(Figure 1).

207

Thorn et al.52 reported a comprehensive study on the chemical properties of both FAs

208

investigated in this work. The results showed the larger aromatic carbon content and C/H

209

ratio (Table 1) in PPFA, which suggest that its aggregates are stabilized to a larger extent by

210

van der Waals forces. Larger content of aromatic carbon systems implies that the amount of

211

π-electrons in PPFA is bigger than in SRFA. π-clouds are responsible for the hyperchromic

212

effect,56 which explains the large HPSEC band observed at ∼ 9 kg mol-1 (Figure 1) and causes

213

the larger PPFA nominal weight average molecular weight (Mw). In addition, PPFA was also

214

shown to have a lower amount of phenolic groups, and a larger content of carboxylic groups

215

as compared to SRFA.52 Under the experimental conditions applied here, most of the

216

carboxylic groups are deprotonated and thus negatively charged. For this reason they repel

217

each other and cause the lower tendency of PPFA (as compared to SRFA) to form inter- and

218

intra-molecular hydrogen bonds,57 thereby supporting the statement about the

219

predominant stabilization by van der Waals forces. On the contrary, SRFA shows lower

220

aromatic carbon content and larger content of phenolic groups.52 Under the experimental

221

conditions used here (i.e. acidic pH), the phenolic groups are protonized and may form H-

222

bonds with the deprotonized carboxylic groups.58 For this reason, we suggest that the H-

223

bonds play more important role in SRFA aggregates stabilization as compared to PPFA.

224

By accepting hypotheses of Drastik et al.32 that at high concentration DOM mainly consists of

225

hydrophobic (i.e. aromatic) core surrounded by hydrophilic moiety formed by progressive

226

aggregation of amphiphilic and polar molecules, we can argue that PPFA consists of large 8 ACS Paragon Plus Environment

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aggregates containing condensed aromatic hydrophobic systems covered by polar, mostly

228

deprotonated carboxylic groups. They allow good solvation and make PPFA more thermally

229

stable and incompressible even upon dilution as also indicated in ref.32 Conversely, the SRFA

230

samples has more hydrogel-like character that allows a better penetration of water

231

molecules and larger structural compressibility, which makes the system more vulnerable to

232

heat-induced transitions.32

233 234

Fast field cycling 1H nuclear magnetic resonance (FFC 1H NMR) relaxometry results

235

The contrasting structural features of both FAs as described above are important for

236

understanding the relaxation of water protons in DOM.

237

FFC NMR experiments revealed a decrease of the R1 values in the whole range of the applied

238

magnetic field (BRLX) for both fulvic acids as temperature was changed from 25 to 52 °C

239

(Figure 2). In particular, all the relaxation rates were faster for the PPFA than for SRFA

240

(compare the y-axes in Figure 2a and 2b).

241

The T1 relaxation mechanism refers to the energy transfer between water and dissolved

242

molecules.55 Two motion regimes, each reflecting different proton longitudinal relaxation

243

mechanisms can occur, as water is located in the close proximity of FAs’ surfaces. The outer-

244

sphere relaxation mechanism (which causes the slow motion regime), refers to water

245

diffusing in rigid structures. As a consequence, the rate of relaxation is reciprocally

246

proportional to the solvent diffusion coefficient.59 The second relaxation mechanism, i.e. the

247

inner sphere one (causing the fast motion regime), is less affected by diffusion, while the

248

chemical exchange between water and the solute (i.e. the fulvic acids) plays the dominant

249

role. Figure 3 shows that the fast motion regime occurs in both FAs.60 However, difference in

250

relaxation rates in both FAs (Figure 2) implies that relaxation of water protons in the

251

hydrogel-like SRFA is slightly decelerated by diffusion. On the contrary, due to its rigidity,

252

PPFA structure is less penetrable and thus diffusion influences relaxation at a lesser extent.

253

To verify this conclusion, the relaxation rates obtained at different temperatures were used

254

to determine activation energies and frequency factors from the slope and y-intercept of the

255

linearized form of the Arrhenius equation (3) (i.e. ln(R1) vs 1000/T), respectively. These

256

parameters were calculated for relaxation rates obtained at Larmor frequencies of 0.08 and

257

30 MHz (Figure 3, and Table 2).



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As reported in Table 2, the activation energies obtained by using R1 values are negative,

259

which resulted from decreasing relaxation rate with increasing temperature. As a rule,

260

negative values of activation energy indicate that the investigated interaction is complex and

261

does not proceed in one step or that the processes proceed along attractive potential energy

262

surfaces without energy barriers on which slower collisions are more reactive than faster

263

ones.61 Therefore, the obtained activation energies should not be simply interpreted in

264

terms of height of the potential (energy) barrier separating two minima. Instead, they are

265

interpreted as values reflecting a complex mechanism of interactions between water and

266

fulvic acids.

267

The second parameter retrieved from the Arrhenius equation is the frequency factor (Table

268

2). This parameter is the factor reflecting the frequency at which the collisions between

269

water and fulvic acids take place regardless their energy and for this reason it is less

270

influenced by interaction complexity. Consequently, the lower frequency factor for SRFA

271

indicates a lower count of interactions between water and SRFA molecules caused by the

272

restricted diffusion of water in SRFA.

273

The activation energy and the frequency factor measured at the proton Larmor frequencies

274

of 30 and 0.08 MHz (Table 2) resulted in different values for SRFA, but similar for PPFA.

275

Measurements at short Larmor frequencies are more sensitive to slowly relaxing pool of

276

water protons, such as the weakly bound or freely moving proton relaxation pools.

277

Conversely, as proton Larmor frequency increases, measurements provide information on

278

the quickly relaxing proton pools, which are due to strong interactions.49 Therefore, the

279

larger span activation energies can be interpreted with respect to FA structure as being

280

related to its heterogeneity. SRFA, due to its hydrogel-like character, contains assumingly

281

more heterogeneous environment for water proton relaxation in its structure than PPFA,

282

where the relaxation between water and polar groups takes place mainly on the surface. For

283

this reason, the values of activation energies are similar at high and low frequencies in PPFA

284

and different in SRFA. This statement accords with the polydispersity index (Table 1), which

285

is a measure of FA heterogeneity, thereby resulting higher for SRFA.

286

The results above explain the peculiarity that although water relaxes faster in PPFA (Figure

287

2), its correlation time (τc) is lower (Figure 4). In fact, correlation time represents a time

288

necessary for rotation of 1 rad of a water molecule.55 In light of above discussion, water

289

rotation is faster in the PPFA than in the SRFA structure (Table 2). Furthermore, Figure 4 10 ACS Paragon Plus Environment

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290

confirms the larger structural rigidity of PPFA caused by its aromatic structure. In fact, Figure

291

4 shows that the correlation times of PPFA surrounding water do not change significantly at

292

different temperatures. In contrast, the τc values of the water system in SRFA decreases with

293

increasing temperature. This means that temperature increment causes an increase in water

294

motion in SRFA due to the weakening of the FA structure. Accordingly, the physical structure

295

of SRFA is less thermally stable as in PPFA, which can be attributed to larger number of H-

296

bond stabilizing SRFA structure. Shaffer and von Wandruszka62 concluded that two

297

counteracting processes are effective at elevated temperatures both thermal agitation,

298

which tends to disrupt the aggregates, and dehydration, which promotes aggregate

299

formation. Therefore, in relatively rigid and aromatic PPFA, the temperature increase causes

300

a disruption of polar assemblies and simultaneously increases hydrophobic effect stabilizing

301

most van der Waals forces63 in hydrophobic assemblies. This leads to the increasing

302

hydrophobicity of hydrophobic domains in diluted humic acids solution upon heating as

303

observed by Shaffer and von Wandruszka62 and indicates that these domains, composed of

304

hydrophobic molecules, are porous, which enables their interaction with hydrophobic

305

compounds. It is also noteworthy that the results in Figure 4 are in contrast to observations

306

using ultrasonic velocimetry, which indicated absence of structural changes with increasing

307

temperature.64

308

Kucerik et al.65 showed that humic and fulvic acids from different sources (IHSS standards)

309

start aggregating already at concentrations around 0.01 g L-1. Zheng and Price66 investigated

310

diffusion coefficients of hydrophilic and aliphatic moieties of DOM and observed their

311

aggregation only at higher concentrations. Similarly, Smejkalova and Piccolo25 reported

312

micelle-like behavior only for two out of nine investigated humic acids and concluded that in

313

the rest of investigated samples, the length of amphiphilic molecules is either too short or

314

their abundance is too low to form micelles. Based on these results, Drastik et al32 concluded

315

that hydrophobic molecules are aggregated already at very low concentration and

316

successively involved in gluing hydrogels as concentration increases. In this work, we showed

317

that structural dynamics of DOM is largely influenced by hydrogel-like structure composed of

318

either amphiphilic or polar molecules stabilized by H-bonds, which is in accordance with

319

results of Lam et al.,36 who observed the importance of carboxyl-rich alicyclic molecules in

320

aggregation of SRFA. This DOM part responses to thermal fluctuation (and other factors such



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321

as pH and ionic strength) and presumably modulates the reactivity of hydrophobic

322

aggregates trapped in the gel structure.

323 324

Structural hysteresis of DOM

325

The relaxation rates R1 values retrieved from the heating and cooling cycles revealed the

326

hysteresis in R1 values for water as well as for both fulvic acids (Figure 5 reports exemplary

327

results only for SRFA). In other words, the rate of water protons relaxation differs as the

328

system is cooled down or heated up during the heating/cooling cycle.

329

Liquid water is a molecular 3D network stabilized by H-bonds and van der Waals forces. The

330

formation of H-bonds brings about energetic changes in enthalpy and entropy of the whole

331

network. Under equilibrium, the enthalpy and the entropy factors are mutually

332

compensated,67 but their response to temperature variation is different. We hypothesize

333

that this difference can cause the delay in re-establishing of water structure. This conclusion

334

is in accordance with the longer relaxation time during water cooling (Figure 5). The slower

335

relaxation rate indicates a change in arrangement of water structure and a decrease in

336

strength of H-bonds. The decreased strength of H-bonds in water influences the self-

337

assembling of dissolved FA molecules and consequently decreases the correlation time of

338

water in FAs upon heating and cooling (Table 2). The correlation times in Table 2 also

339

indicate that the structural hysteresis is more pronounced in hydrogel-like SRFA, i.e. in

340

flexible system containing more H-bonds than in rigid, aromatic PPFA. The results in Figure 5

341

suggest that delay in re-conformation of 3D water network is primary reason of re-

342

establishing of H-bonds in FA structure. This is in accordance with recent conclusions that

343

break of H-bonds in diluted FA and humic acids solutions are the main reasons of phase

344

transitions at 20 and 40°C and in shift of their temperatures upon heating and cooling.32

345

The variation of temperature in nature is a rule. Investigation of water properties in two

346

structurally contrasting FA showed that the temperature variation changes the properties of

347

water structure, thereby changing the properties of self-assembled colloidal systems. The

348

character of DOM self-assemblies influences their reactivity and mobility, which indicates

349

that partitioning of DOM with pollutants and nutrients and their transport depend on DOM

350

thermal history. Future research is needed in order to reveal the factors influencing the

351

DOM hysteresis. In addition, a new question arises: how relevant is DOM hysteresis in


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ecological processes such as the non-reversible sorption of and the fate of engineered

353

nanoparticles, nutrients and contaminants.

354 355

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503 504 505

Captions to Figures

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(dotted line) fulvic acids.

507

Figure 2. Nuclear magnetic resonance dispersion (NMRD) profiles of the Pahokee peat (a)

508

and Suwannee river (b) fulvic acids. The continuous lines are obtained by the fitting

509

procedure described in the text.

510

Figure 3. Relationship between the longitudinal relaxation times measured at 0.08 and 30

511

MHz for the Pahokee peat (a) and Suwannee river (b) fulvic acids.

512

Figure 4. Variation of the water correlation time (tC) during the heating cycle of the solutions

513

containing Suwannee river (black line) and Pahokee peat fulvic acids (red line).

514

Figure 5. Hysteresis in longitudinal relaxation time during the heating and cooling cycle of

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demineralized water (a) and Suwannee river fulvic acid (b).

Figure 1. HPSEC chromatograms of the Suwannee River (continous line) and Pahokee Peat

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523 524 525

526 527 528 529 530

531 532 533 534


Table 1. Selected properties of IHSS fulvic acids standards obtained from http://www.humicsubstances.org/ and Thorn et al.52 C H N O C/H C/O Carboxylic Phenolic C/(O+N) Suwannee river 53.0 4.4 0.8 43.9 12.2 1.20 1.19 12.2 3.11 Pahokee peat 52.1 3.2 2.4 43.9 16.1 1.18 1.12 15.2 1.78

Table 2. Parameters determined from HPSEC and FFC NMR measurements Averaged molecular weight Activation energy (kDa) (kJ/mol) Weight Number PDI** 30 MHz* 0.08 MHz* (Mw) (MN) SRFA 9.03 7.20 1.25 18.1±1.1 25.9±1.2 PPFA 9.53 8.29 1.14 11.0±0.8 10.6±0.4 *Larmor frequency, ** polydispersity index

Frequency factor (s-1) 30 MHz*

0.08 MHz*

(2.9±0.2)×10-4 (9.8±0.2)×10-3

(1.3±0.1)×10-6 (1.5±0.1)×10-2

Correlation time (ns) Before After heating heating 15 2 0.28 0.19

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Figure 1. HPSEC chromatograms of the Suwannee River (continous line) and Pahokee Peat (dotted line) fulvic acids. 271x192mm (300 x 300 DPI)


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Figure 2a. Nuclear magnetic resonance dispersion (NMRD) profiles of the Pahokee peat (a) and Suwannee river (b) fulvic acids. The continuous lines are obtained by the fitting procedure described in the text. 272x221mm (300 x 300 DPI)



Figure 2b. Nuclear magnetic resonance dispersion (NMRD) profiles of the Pahokee peat (a) and Suwannee river (b) fulvic acids. The continuous lines are obtained by the fitting procedure described in the text. 271x220mm (300 x 300 DPI)


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Figure 3. Relationship between the longitudinal relaxation times measured at 0.08 and 30 MHz for the Pahokee peat (a) and Suwannee river (b) fulvic acids. 366x276mm (237 x 237 DPI)



Figure 4. Variation of the water correlation time (tC) during the heating cycle of the solutions containing Suwannee river (black line) and Pahokee peat fulvic acids (red line). 387x266mm (226 x 226 DPI)


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Figure 5. Hysteresis in longitudinal relaxation time during the heating and cooling cycle of demineralized water (a) and Suwannee river fulvic acid (b). 361x465mm (300 x 300 DPI)



graphical abstract 40x27mm (300 x 300 DPI)


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Water Dynamics and Its Role in Structural Hysteresis of Dissolved

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