Development of analytical methods for ammonium determination in seawater over the last two decades

Journal Pre-proof Development of analytical methods for ammonium determination in seawater over the last two decades Yong Zhu, Jianfang Chen, Dongxing Yuan, Zhi Yang, Xiaolai Shi, Hongliang Li, Haiyan Jin, Lihua Ran PII:

S0165-9936(19)30409-1

DOI:

https://doi.org/10.1016/j.trac.2019.115627

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TRAC 115627

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Trends in Analytical Chemistry

Received Date: 12 July 2019 Revised Date:

9 August 2019

Accepted Date: 9 August 2019

Please cite this article as: Y. Zhu, J. Chen, D. Yuan, Z. Yang, X. Shi, H. Li, H. Jin, L. Ran, Development of analytical methods for ammonium determination in seawater over the last two decades, Trends in Analytical Chemistry, https://doi.org/10.1016/j.trac.2019.115627. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2019 Published by Elsevier B.V.

Development of analytical methods for ammonium determination

1 2

in seawater over the last two decades

3

Yong Zhua, Jianfang Chen*a, Dongxing Yuanb,a, Zhi Yanga, Xiaolai Shia, Hongliang Lia,

4

Haiyan Jina, Lihua Rana

5

a

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Administration & Second Institute of Oceanography, Ministry of Natural Resources, P. R.

7

China

8

b

9

and Ecology, Xiamen University, Xiamen 361102, P. R. China

Key Laboratory of Marine Ecosystem and Biogeochemistry, State Oceanic

State Key Laboratory of Marine Environmental Science, College of the Environment

10

*

11

E-mail: [email protected]

12

Address: No. 36 Baochubei Road, Xihu District, Hangzhou 310012, Zhejiang Province, P.

13

R. China

Corresponding author

14 15

Abstract

16

Ammonium plays an important role in the nitrogen cycle in marine environments.

17

There is no universal method for ammonium analysis that can be applied to all types of

18

seawater and meet the requirements of different applications. Therefore, selecting the

19

most appropriate method is of crucial importance. The aim of this review is to explore the

20

diverse range of methods available for the detection and analysis of ammonium in

21

seawater, to provide a basis for selection of the most suitable method. The developments

22

of typically used methodologies for the analysis of seawater are summarized, including 1

23

ion-selective electrode, spectrophotometric, fluorometric and matrix separation methods.

24

The main parameters assessed in research published in the last two decades (1999 to 2019)

25

are also reviewed. To make this review specific to seawater analysis, only methods

26

dealing with actual coastal, estuarine and seawater analysis or having specific salinity

27

effect evaluations, were selected.

28

Keywords: Ammonium; Ammonia; Marine environment; Flow analysis; Ion-selective

29

electrode; Spectrophotometric; Fluorometric; Matrix separation; Ammonium-free

30

seawater

31 32

1. Introduction

33

Nitrogen is a limiting element for biological productivity, occupying a central role in

34

marine biogeochemistry. The marine nitrogen cycle is one of the most complex marine

35

biogeochemical cycles, as more chemical forms of nitrogen exist (nine possible oxidation

36

states) than most other elements, with a myriad of potential chemical transformations[1].

37

Among all nitrogen species present in seawater environments, ammonium is the most

38

reduced form of inorganic nitrogen, and is the preferred form of nitrogen for marine

39

phytoplankton. Therefore, ammonium plays an important role in nitrogen cycling in

40

marine environments, especially in the euphotic zone of oligotrophic regions. The

41

ammonium ion (NH4+) is the dominant form, while ammonia (NH3) is a minor component

42

in marine environments and therefore, the sum of ammonia and ammonium ions is

43

referred to as ammonium in this paper. The dissociation equation of ammonium ion to

44

ammonia[2] is shown as 2

45 46

The relative flow of ammonium via assimilation by phytoplankton versus oxidation by

47

microbial nitrifiers, largely determines the composition of the upper ocean nitrogen pool

48

and therefore, plays a crucial role in controlling marine productivity and the export of

49

fixed carbon throughout the ocean[3]. The ammonium concentration reflects the balance

50

between production via ammonification and consumption via ammonium assimilation or

51

nitrification. Furthermore, ammonium concentrations provide valuable insight into marine

52

nitrogen transformations, such as ammonium assimilation, nitrification and anammox

53

processes. Establishing an accurate ammonium concentration is beneficial for

54

characterization of water masses when modeling nutrient fluxes and estimating algal

55

growth potential[4]. Additionally, knowledge of ammonium concentrations in seawater

56

also furthers our understanding of the underlying regulatory mechanisms of marine

57

nitrogen processes.

58

Ammonium is considered to be one of the most important pollutants in aquatic

59

ecosystems, inducing ecological stress and serving as a key indicator of water quality [5].

60

The determination of ammonium concentrations and fluxes, along with a comprehensive

61

understanding of the complex biogeochemical processes, is imperative for the effective

62

management and conservation of sustainable marine ecosystems and their catchment

63

areas[6]. Higher concentrations of ammonium, especially the more toxic form ammonia,

64

has been shown be harmful to aquatic organisms such as fish, shrimp, abalone, and sea

65

urchins, with the highest sensitivity to exposure during larval or juvenile development

66

stages[7]. In most natural water environments, ammonium exists mainly in the form of 3

67

ammonium ions, while ammonia becomes the dominant species when the pH increases to

68

above 9.75. Both forms are easily interconverted, with the ratio of ammonia to

69

ammonium ions largely depending on pH and temperature[5]. Additionally, ammonium

70

ions have also been found to contribute to toxicity as they interfere with the outward

71

movement of ammonia via the gills[8]. Large amounts of ammonium can enter the

72

aquatic environment from anthropogenic sources such as municipal effluent discharges

73

and agricultural runoff. In addition, ammonium is supplied by natural sources such as the

74

excretion of nitrogenous waste from animals, contributing to eutrophication, the

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development of harmful algal blooms, decreased oxygen levels and death of biota in

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aquatic environments[9]. The risk of ammonium stress is increased in intensive

77

aquaculture environments and therefore, accurate monitoring of ammonium concentration

78

is necessary for the evaluation of water quality and the protection of aquaculture stocks.

79

Furthermore, ammonium monitoring is essential to further our understanding of nitrogen

80

cycling in aquatic ecosystems.

81

Accurate measurement of ammonium seems to be the most challenging in seawater

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nutrient analysis, although various methods for ammonium analysis have been proposed.

83

Several comprehensive reviews of ammonium detection methods for seawater analysis

84

have been performed. Šraj et al.[6] reviewed the analytical challenges and advantages of

85

using flow-based methodologies for ammonium determination in estuarine and marine

86

waters, with discussion of detection and on-line sample-pretreatment strategies to

87

improve the limit of detection and to reduce or eliminate interferences, with field

88

application examples. Ma et al.[10] reviewed procedures for analysis of nutrients 4

89

including ammonium at nanomolar levels in seawater, with aspects of measurement

90

protocols that affect the quality of analyses of trace nutrients summarized, including

91

contamination of reagents, sample storage, and the preparation of nutrient-free seawater.

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In addition, Ma et al. [11] also reviewed the applications of flow techniques in seawater

93

analysis, with the determination of ammonium from 2008 to 2015. However, the

94

aforementioned studies by Šraj et al.[6] and Ma et al. [10, 11] all focused on flow-based

95

methods for ammonium analysis or methods for nanomolar level ammonium

96

measurements. Considering the large number of available methods, selection of a suitable

97

method is difficult as no single method that can be applied universally to all types of

98

seawater, while meeting all the requirements of different applications. Selecting the most

99

appropriate method for ammonium analysis is important as it determines the cost of

100

analysis in terms of operational complexity and instrumental effort. This selection is also

101

influenced by framework conditions, such as sample size, duration of analysis, sample

102

availability and prior information about the sample contents[12]. Therefore, a review of

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the different available methods for ammonium measurement in seawater is urgently

104

needed to help analysts select the optimum method according to the specific conditions

105

and requirements. In 2006, Molins-Legua et al.[5] created a guide for selection of the

106

most appropriate method for ammonium determination in water, by critically evaluating

107

the main parameters involved in determining ammonium in water samples. The evaluated

108

methods included the Nessler method, ion-selective electrodes, indophenol-type reagents,

109

Roth’s fluorometry and methods based on the measurement of chemiluminescence

5

110

generated by luminol and TCPO reagents. However, while many methods are suitable for

111

aqueous samples, few can be applied to the more complex matrix of seawater.

112

The aim of this review is to explore the diverse range of methods available for

113

ammonium detection and analysis in seawater, to assist selection of the most suitable

114

method. The developments of typically used methodologies in seawater analysis are

115

summarized including ion-selective electrode, spectrophotometric, fluorometric and

116

matrix separation methods. Furthermore, the main parameters assessed in the studies

117

published in the last two decades (1999 to 2019) are also reviewed. To ensure this review

118

is more specific for seawater analysis, only methods dealing with actual coastal, estuarine

119

and seawater analysis or evaluating specific salinity effects were selected.

120

2. Methodologies for ammonium analysis

121

2.1 Ion-selective electrode method

122

There is a significant need for the collection of chemical data in natural water systems

123

at a high temporal and spatial resolution using in-situ measurements, to eliminate biases

124

resulting from the preservation or storage of samples and the alteration of environmental

125

conditions. The ion-selective electrode (ISE) method provides the benefit of in-situ

126

analysis with the additional advantages of easy operation, fast response times,

127

miniaturized size, low power consumption, low manufacturing costs, and a wide dynamic

128

response range. Therefore, ISE has become an attractive sensing platform for

129

environmental water analysis[13].

130 131

ISE is also referred to as a specific ion electrode, as it responds to the concentration of a particular ion or gas in solution. Two types of electrodes can be used for ammonium 6

132

measurement in natural water[14]. One is ammonia gas sensing probe using a hydrophilic

133

gas-permeable membrane (typically PTFE) to separate ammonia from the sample with an

134

internal solution of ammonium chloride[12]. Since in most natural waters ammonium

135

exists mainly as ammonium ion, it is necessary to convert all ammonium ions to ammonia

136

by adding a strong base, such as lithium hydroxide. Ammonia diffuses through the

137

membrane until the partial pressure of ammonia is equal on both sides, generating a

138

potential difference that can be measured using a high impedance voltmeter (i.e. pH

139

meter). Another one is ammonium ion sensing electrode with a polyvinylchloride (PVC)

140

membrane containing an ammonium-carrier. The water sample is acidified to lower the

141

pH and convert essentially all ammonia to ammonium ions. The electrode potential of the

142

sample relative to the reference electrode of the ammonium ion sensing probe is

143

proportional to the ammonium ion concentration in the sample[14]. Chen et al.[15]

144

summarized the use of electronic sensors (including potentiometric, voltammetric and

145

field-effect transistor sensors) and their performance for nitrate, nitrite, ammonium and

146

phosphate detection in an aqueous environment. While Crespo[13] presented recent

147

advances in polymeric-based ISEs relevant to water research, with the challenges of ISE

148

application in saline water also discussed.

149

Gilbert and Clay[16] investigated the determination of ammonium in seawater using an

150

ammonia gas sensing probe (Orion ammonia electrode model 95-10) in 1973. Seawater

151

samples were successfully measured using this method, although extremely long

152

equilibration times were required when the concentration of ammonium was below 7.1

153

µM. Additionally, they indicated that measurement of ammonium in samples with varying 7

154

concentrations of other potentially interfering ions, should be carried out according to the

155

well-established addition method. Merks[17] modified the Gilbert and Clay[16] method,

156

with the practicality of use assessed in marine and estuarine water samples. With salinity

157

influence correction, this method proved to be reliable for concentrations over 7.1 µM, to

158

the highest tested concentration of 142.9 µM. Moschou et al.[18] developed a novel

159

portable flow analysis system using ion-selective electrodes as detectors, allowing direct

160

electrochemical ammonium and nitrite monitoring for the simultaneous measurement of

161

ammonium and nitrite in seawater and aqua-culture samples. The system was capable of

162

suitable operation in non-filtered samples such as marine aquaculture rearing medium and

163

wastewater treatment plant effluents within the ammonium and nitrite concentration range

164

of 3.6-714.3 µM. Wen et al.[19] developed a real-time and reagent-free method using an

165

ammonium ion-selective electrode for the real-time measurement of ammonium, with

166

compensation for the ammonium ion component of the total ammonium fraction. Results

167

showed that the ammonium ion ratio in ammonium could be calculated based on pH and

168

temperature within the ammonium concentration range of 7.1-714.3 µM. However, K+

169

cations were found to interfere with the measured potential of the ammonium ISE,

170

especially at low ammonium concentrations. Therefore, compensation is necessary when

171

measuring solutions containing K+.

172

ISE methods for ammonium have been applied to ammonium detection at

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concentration ranging from 3.6 to 714.3 µM, with the inherent poor sensitivity of ISE

174

limiting its widespread application. To improve the sensitivity of ISE, carbon nanotube

175

based ion sensors[20], solid-contact potentiometric sensors[21] and some indirect 8

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electrochemical methods[22] have been developed. A carbon nanotube based sensor

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insulated using a photoresist film was developed by Jang et al.[20] for the measurement

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ammonium in artificial seawater, which expose only the carbon nanotube sensing section

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of the probe, allowing the successful detection of a 10 nM ammonium solution ~pH 6.

180

Ding et al.[21] developed a solid-contact potentiometric sensor based on a solid-contact

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ammonium selective electrode for in-situ detection of ammonium in seawater. The

182

all-solid-state polymeric membrane ammonium-selective electrode was integrated with a

183

polyvinyl alcohol hydrogel buffer film at pH 7.0, with a gas-permeable membrane. The

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ammonia in seawater diffused through the gas-permeable membrane and was converted to

185

ammonium ion in the hydrogel buffer, which could be potentiometric ally sensed by the

186

solid-contact ammonium-sensitive membrane electrode, showing a detection limit of 0.64

187

µM with a linear range of 1-100 µM. Based on the reaction of HBrO with ammonium, an

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indirect electrochemical method was proposed by Takahashi et al.[22] for the

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determination of ammonium in a phosphate buffered solution at pH 7 using a dual

190

electrode configuration. In this system, HBrO was produced at a generator electrode and

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the excess HBrO was subsequently detected at a collector electrode after reaction with

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ammonium, resulting in a detection limit of below 3.0 µM.

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2.2 Spectrophotometric method

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Spectrophotometry is the most commonly used method for ammonium determination

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in aqueous systems, as well as the adopted standard method of the U.S. Environmental

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Protection Agency and other national testing agencies. The spectrophotometric methods

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applied for ammonium measurement include the Nessler’s reagent method (or 9

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Nesslerization method), the indophenol blue (IPB) method, other IPB type methods and

199

hypobromite oxidation spectrophotometry. In addition, a spectrophotometric method

200

based on the reaction of ammonium, o-phthaldialdehyde (OPA) and sulfite has been

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applied, which is normally used in fluorometric analysis. Nessler’s reagent is a solution

202

consisting of mercury (Ⅱ) iodide and potassium iodide in a highly alkaline solution,

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which reacts with ammonium resulting in a yellow-brownish complex, allowing

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colorimetric determination of the amount of ammonium present[14]. However, Nessler’s

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reagent easily reacts with calcium and magnesium causing precipitation or turbidity that

206

interferes with colorimetric measurements[14]. To avoid matrix interference, sample

207

pretreatment methods such as distillation, are often required to separate ammonium from

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the sample matrix[23]. In addition, the toxic reagent mercury is used in the Nessler’s

209

reagent method, which presents the risk of secondary pollution and significantly limits the

210

applicability of this method[12]. Furthermore, the Nessler’s reagent method is often used

211

in wastewater or freshwater analysis and has seldom been applied to seawater analysis as

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the method has been found to have an unsatisfactory performance in moderately hard

213

waters (e.g. seawater).

214

The IPB method based on the Berthelot reaction is the most widely used colorimetric

215

method for the determination of ammonium in seawater. The reaction of hypochlorite,

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alkaline phenol solution and ammonium, forming an indophenol blue dye, was first

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reported by Berthelot in 1859[24] and since then a large number of modifications have

218

been made to the basic reaction. Searle[23] performed a comprehensive review of the

219

development of IPB methods in 1984 including the historical development of methods, 10

220

the mechanism of the reaction, reaction conditions (e.g. pH, reagent concentrations,

221

reaction time, reaction temperature and sequence of addition of reagents), interferences,

222

and applications. Modifications to the reaction mechanism and reagents are mainly

223

associated with the selection of suitable reagents: 1) phenolic compound or other

224

substitute reagent; 2) hypohalite source; 3) catalyst; 4) chelating agent.

225

In the classic IPB method, phenol is generally used as a reagent. However, other

226

phenolic compounds include thymol, salicylic acid (and its salts), 1-naphthol[25],

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guaiacol, o-phenylphenol (OPP), o-chlorophenol, 2-methyl-5-hydroxyquinoline and

228

m-cresol have also been previously used for IPB dye formation as alternatives to

229

phenol[23]. The use of thymol appears to be limited mainly to solvent extraction

230

methods[23], while the use of other phenolic compounds (except sodium salicylate and

231

OPP) have been also seldom applied in more recent studies. Different IPB methods using

232

phenol, salicylate and OPP will be described in the section 2.2.3.

233

Hypochlorite is another important reagent in the IPB method, with hypobromite,

234

chloramine-T and sodium dichloroisocyanurate (DIC) also used in the reaction as

235

alternatives. DIC has been proven to be both more convenient to use and more stable in

236

solution, than sodium hydrochlorite. However, DIC may not be suitable for some

237

applications as it reacts with protein and amines, causing reaction interference by

238

lowering the effective hypochlorite concentration. Thus, hypochlorite or DIC are the most

239

widely used reagents in recent studies.

240 241

To increase the speed of reaction in the IPB method, a high reaction temperature and a catalyst are required. Catalysts such as manganese (Ⅱ) ion, acetone, sodium nitroprusside 11

242

(sodium nitroferricyanide), sodium pentacyanonitrosylferate, sodium

243

aquopentacyanoferrate and potassium hexacyanoferrate (Ⅱ), have been used to accelerate

244

the formation of indophenol. Since the 1960’s, the nitroprusside-catalyzed reaction has

245

become increasingly popular and is the most widely reported method for measurement of

246

ammonium in solution.

247

To increase the sensitivity of the reaction in seawater a high pH is required, which is

248

usually achieved by increasing the final sodium hydroxide concentration, resulting in the

249

precipitation of hydroxides from seawater. To overcome the interference of precipitation

250

and minimize the effects of variations in salinity in the IPB method, a chelating agent

251

such as tartrate, trisodium citrate, citrate, 1,2-cyclohexane diamine tetraacetic acid

252

(CDTA), or ethylene diamine tetraacetic acid (EDTA) is typically used as complexing

253

agent.

254

The flow analysis concept emerged during the 1950s, with the advent of segmented flow

255

analysis (SFA). Since then, the concept has evolved and many methodologies for

256

automated ammonium measurement have been widely applied in seawater analysis.

257

Flow-based methodologies have the advantage of not only permitting automated analysis,

258

but also improving sensitivity and reproducibility, while providing the possibility of

259

miniaturization or portability. This allows for remote field deployment, providing high

260

quality analytical data with good temporal and spatial resolution. It has been established

261

that flow-based methods generally produce better results for ammonium monitoring, than

262

manual methods[26]. Šraj et al.[6] performed a comprehensive review of the available

263

flow-based methodologies for ammonium determination. However, even with flow 12

264

techniques, the sensitivity of traditional spectrophotometric methods remains unable to

265

meet the requirements of low level ammonium measurements. The ammonium

266

concentrations reported in seawater worldwide, especially in oligotrophic oceans and

267

deep oceans, are typically below 1 µM[27]. According to a review on the determination of

268

nanomolar levels nutrients in seawater[10], two approaches are typically used to improve

269

the sensitivity of spectrophotometric methods, either by preconcentrating analytes or

270

analyte-derivatives, or by increasing the path-length of the absorption cell.

271

2.2.1 IPB method using phenol

272

In the classic IPB method, phenol, hypochlorite and nitroprusside are generally used as

273

regents. Under alkaline conditions, ammonia reacts with hypochlorite to form

274

monochloramine, which then reacts with two molecules of phenol to form blue-colored

275

indophenol. Schemes of the chemical reaction are shown in Fig. 1.

276 277 278

Fig. 1 Schemes of the chemical reaction of the IPB method using phenol

279

Aminot et al.[26] reported the 5th ICES intercomparison exercise for nutrients in

280

seawater. Disparities in the ammonium results submitted from 106 laboratories (standard

281

deviations of 22-23% at medium and high concentrations and 56% at low levels) showed

282

that accurate determination of ammonium remains a problem in the oceanographic

283

community. The drawbacks of the IPB method using phenol include the use of toxic

284

reagents, high levels of blank, low sensitivity and interference from seawater matrices.

285

Some of these problems have been solved by modification of the basic IPB method. The 13

286

pH shift problem in seawater analysis using the IPB method was investigated by Pai et

287

al.[28], with hydrolysis of the magnesium-citrate complex found to be the main cause of

288

decreased pH in the final sample solution, requiring more sodium hydroxide to overcome

289

the buffering capacity of seawater. Using the gas diffusion (GD) technique allows

290

separation of ammonium from the sample solution and the elimination of matrix effects,

291

which will be discussed in the section 2.4.1. In order to improve the sensitivity of the IPB

292

method using phenol, several approaches have been proposed for preconcentration of

293

ammonium or its derivatives.

294

Such as solvent extraction, solid phase extraction (SPE), headspace single-drop

295

microextraction (HS-SDME), micro-phase sorbent extraction, membrane filtration (MF),

296

Amberlite XAD-7, mixed micelle-mediated extraction (mixed-MME) and liquid phase

297

microextraction (LPME). These preconcentration techniques have been coupled with the

298

IPB method and successfully applied to the determination of ammonium in environmental

299

samples. However, few of these methods have been used for ammonium measurement in

300

seawater. A solvent extraction method for nanomolar ammonium measurement in

301

seawater, was described by Brzezinkski[29]. The IPB dye formed by reaction of phenol,

302

hypochlorite and ammonium using sodium aquopentacyanoferrate as a coupling agent

303

was concentrated by extraction into n-hexanol at a low pH, followed by re-extraction into

304

an aqueous alkaline buffer. Clark et al.[30] developed a sensitive method based on the

305

IPB method, combining the advantages of ammonium-specific derivatization and

306

preconcentration of ammonium by SPE. The IPB dye was collected by SPE using a C18

307

cartridge and eluted by methanol, with the IPB containing eluent then assessed by 14

308

GC-MS. Another method combining SPE, sequential injection analysis (SIA) and the IPB

309

method using phenol was proposed for trace ammonium measurements by Chen et al.[31].

310

The formed IPB compound was extracted onto a hydrophilic-lipophilic balance (HLB)

311

cartridge and eluted using a solution containing 30% (v/v) ethanol and 5.0 mM

312

sodium hydroxide, with determination by spectrophotometry at 640 nm. Under the

313

optimized conditions, an LOD of 3.5 nM was obtained with a linear range of 0-428 nM,

314

while the salinity effect was ignored. However, the sample throughput of this method was

315

only 3 h-1 and it has not been applied to field-based determination of ammonium in

316

seawater.

317

Another approach used to improve the sensitivity of colorimetric methods is increasing

318

the path-length of the absorption cell. According to the Lambert-Beer law, the absorbance

319

of a sample increases with extension of the optical path length, thus enhancing the

320

sensitivity[10]. Li et al.[32] developed an automated system for nanomolar level

321

ammonium determination in seawater by spectrophotometry, using SFA coupled with a

322

long path liquid waveguide capillary cell (LWCC). Phenol, DIC and sodium

323

nitroferricyanide were used as the main reagents, with a mixture of citrate and EDTA

324

added as a complexing agent to prevent precipitation. The optimal concentration of the

325

reagents and parameters of the flow system were discussed, with a LOD of 5 nM obtained.

326

With the addition of an auto-sampler system, the method was applied in Florida Bay and

327

Biscayne Bay for analysis of the distribution of ammonium. Zhu et al.[33] also developed

328

an automated colorimetric method for the on-line determination of trace ammonium in

329

seawater, using the flow injection analysis (FIA) technique coupled with a LWCC. The 15

330

same reagents and complexing agent were used in this method as outlined by Li et al.[32],

331

with low reagent consumption noted to reduce the use of toxic reagents and the risk of

332

secondary pollution. Under optimal conditions, this method provided an LOD of 3.5 nM

333

with a linear range of 10-300 nM, although the linear range could be extended by

334

choosing a less sensitive detection wavelength. Using this method, the salinity effect was

335

negligible and the Schlieren (refractive index) effect was also found to be negligible if the

336

salinity of the sample was higher than 21. This method was applied in-field for the 24 h

337

on-line monitoring of ammonium in Wuyuan Bay and used to analyze the surface

338

seawater samples collected from the South China Sea. By combining a continuous SFA

339

system, with the GD technique and LWCC, Kodama et al.[34] developed a highly

340

sensitive ammonium determination method, which was found to be accurate over a wide

341

range of concentrations and largely independent of salinity effects, with a LOD of 5.5±1.8

342

nM and linear calibration up to 2000 nM. In addition, the ammonium concentrations were

343

examined in a range of matrices, including ultrapure water, ion-exchanged water, artificial

344

seawater, unfiltered low nutrient seawater (LNSW), filtered LNSW and alkaline LNSW.

345

The method was applied in the vicinity of the Kuroshio Current along 138°E (the O-line)

346

in summer and the typical vertical profiles for ammonium concentration were obtained.

347

2.2.2 IPB method using salicylate

348

Phenol is the most commonly used reagent in the IPB method, presenting operational

349

difficulties as it is caustic, odorous, toxic and exists in transition between solid and liquid

350

phases at room temperature[35, 36]. Therefore, salicylate has been proposed as a phenol

351

alternative for IPB reaction, providing the advantages of lower toxicity and being easier 16

352

to prepare than phenol. Schemes of the chemical reaction of the IPB method using

353

salicylate are shown in Fig. 2.

354 355

Fig. 2 Schemes of the chemical reaction of the IPB method using salicylate The IPB method using salicylate was established after the classic phenate method, with

356

major modifications made, particularly in early studies. Kempers and Kok[37]

357

re-examined the IPB method using salicylate, with optimization of the concentration,

358

preparation and timing of addition of reagents, as well as the reaction temperature and

359

protection from light exposure, resulting in a LOD of 0.4 µM. Muraki et al.[38]

360

developed an automated system for the continuous monitoring of ammonium in a

361

seawater aquaculture environment, using an FIA technique based on the reaction of IPB

362

using salicylate, with the pH of the samples adjusted to 6-7 in advance to avoid

363

precipitation. Jüttner[39] compared the interference of other naturally occurring

364

nitrogen-containing compounds in ammonium determination via IPB with salicylate/DIC

365

and phenol/hypochlorite, finding that the IPB method with salicylate/DIC showed strong

366

interference from all amino acids and peptides tested. In addition, the reactivity of

367

salicylate is lower than that of phenol due to the presence of a carboxy group and

368

therefore, a large amount of salicylate is required to obtain the same level of

369

sensitivity[37]. It is of note, that the use higher pH for optimum color development has

370

largely been overlooked. In 2012, Le and Boyd[40] performed a comparison of phenate

371

and salicylate IPB methods for the determination of ammonium in freshwater and saline

372

water. Results showed that the salicylate method had better precision and accuracy than

373

the phenate method, exhibiting highly satisfactory results using the salicylate method with 17

374

salinities ranging from 7.1 to 1714 mM. In a recent study by Zhou and Boyd[14], the

375

determination of ammonium in aquaculture was performed with comparison between the

376

Nessler’s reagent, phenate, salicylate and ISE methods. The salicylate method was found

377

to be more suitable for application in aquaculture as no hazardous secondary pollution is

378

generated, with high precision and accuracy achieved. In conclusion, although the IPB

379

method using salicylate has been widely applied in environmental and wastewater

380

monitoring, the use of this method in seawater or saline water applications few reports in

381

recent literatures.

382

2.2.3 IPB method using OPP

383

The use of o-phenylphenol (OPP) has also been investigated as a substitute for phenol

384

in the IPB reaction, as it is available in a stable solid state (tabular flaky crystals) and has

385

no causticity, odor, and lower toxicity compared to phenol. In addition, the OPP-based

386

indophenolic compound is not significantly affected by amino acids or urea under a

387

relatively large salinity range[41]. Several studies have reported the use of the IPB

388

method with OPP for application in seawater ammonium analysis, especially in recent

389

years. A possible reaction pathway of the IPB method using OPP was described by Ma et

390

al.[36], as shown in Fig. 3.

391 392 393

Fig. 3 Schemes of the chemical reaction of the IPB method using OPP OPP was first introduced as an alternative to phenol in the IPB method in 1968[41].

394

Kanda[41] then applied the IPB method using OPP for ammonium determination in

395

seawater with monitoring at an absorption wavelength of 670 nm, finding no significantly 18

396

interference by the presence of amino acids and urea, or over a wide salinity range

397

(43-100% seawater). Based on the IPB reaction with OPP, Hashihama et al.[35] described

398

a highly sensitive colorimetric method for the determination of nanomolar concentrations

399

of ammonium in seawater, using a gas-segmented continuous flow analyzer equipped

400

with a long path LWCC and UltraPath, achieving a LOD of 4 nM with a linear range up

401

to 200 nM. This analytical system was applied to underway surface monitoring and

402

vertical observations in the oligotrophic South Pacific, to investigate the distribution of

403

nanomolar ammonium. Lin et al. [42] developed an automated method for the

404

determination of ammonium in estuarine and coastal waters, using reverse flow injection

405

analysis based on the IPB method with OPP, achieving a LOD of 0.08 µM with a linear

406

range up to 35 µM in seawater. The salinity effect was carefully investigated, showing

407

that calibration curve salinity correction was required for the determination of ammonium

408

concentrations. The method was applied to 24 h on-line analysis of ammonium in coastal

409

areas, with a sample throughput of 30 h-1. Ma et al.[36] reported a comprehensive study

410

of the reaction kinetics of IPB with OPP, under different reagent concentrations, reaction

411

temperatures and salinity levels, investigating the salinity interference effects and reagent

412

storage requirements. The reported optimized method allowed manual determination of

413

ammonium in the routine analysis of both freshwater and seawater samples, with no

414

salinity interference and therefore, no need to correct for salinity effects. Under the

415

optimized conditions, this method provided a LOD of 0.2 µM with a linear range up to

416

100 µM. Following this, Ma et al.[43] developed an automated integrated

417

syringe-pump-based environmental water analyzer (iSEA) based on a flow-bath system, 19

418

with a syringe pump used to overcome the drawbacks of batch and continuous flow

419

analyzers. Based on the IPB method using OPP, the automated iSEA was applied to the

420

continuous real-time monitoring of ammonium variations in a river for 24 h and 14 days

421

with a sample throughput of 12 h-1. This fully automated analyzer achieved a detection

422

limit of 0.12 µM, with linear calibration range of 0-20 µM and 0-70 µM with detection at

423

700 nm and 600 nm, respectively. Furthermore, after optimization in both pure water and

424

seawater matrices, the iSEA system was combined with an on-line filtration system for

425

underway analysis of ammonium in coastal areas during 7 cruises[44].

426

2.2.4 Spectrophotometry based on the reaction of NH3-OPA-sulfite

427

The reaction of ammonium with OPA has been well established for fluorescence

428

measurement of ammonium, with Goyal et al.[45] first reporting the feasibility of using

429

spectrophotometry to determine the product of ammonium and OPA for ammonium

430

analysis. Recently, Liang et al.[46] established a spectrophotometric method using FIA

431

for ammonium determination based on the reaction of ammonium, OPA and sulfite as

432

measured at 550 nm. The reaction product generated was rose red at pH > 10.4, with a

433

LOD of 7 µM achieved and no significant difference found between the results obtained

434

from this method and the classical IPB method. The method was applied to the

435

determination of ammonium in lake water, river water, groundwater and sewage.

436

2.2.5 Hypobromite oxidation spectrophotometry

437

The indirect spectrophotometric method of hypobromite oxidation spectrophotometry

438

has also been commonly used for ammonium measurement in seawater. However, this

439

method has often been overlooked in recent studies and reviews. The principle of this 20

440

method is the use of a strong oxidation agent to convert ammonium to nitrite, with nitrite

441

determined via the Griess method. Richards and Kletsch[47] employed the oxidation of

442

ammonium to nitrite by reaction with hypochlorite in the presence of potassium bromide.

443

The method was modified by Matsunaga and Nishimura[48], where the oxidation

444

duration was reduced to 2 min and the interference of amino acids was eliminated. Using

445

this modified method, analysis of various seawater samples from coastal regions showed

446

good correlation with results generated using the IPB method. Liu et al.[49] developed an

447

automated method coupling the reaction with reversed flow injection technique (rFIA),

448

allowing the interference of nitrite and sample turbidity to be removed. Minamiya et

449

al.[50] employed SPE cartridge pre-treatment with hypobromite oxidation

450

spectrophotometry to improve the sensitivity, achieving a LOD of below 38 nM. Sun et

451

al.[51] developed a fully automated spectrophotometric analyzer for the continuous,

452

automated determination of inorganic nitrogen. However, the analyzer was only applied

453

to the determination of standards and artificial samples and not applied in natural water.

454

Tovar et al.[52] employed an automated hypobromite oxidation method with FIA for the

455

speciation of dissolved inorganic nitrogen in seawater, using hypochlorite and potassium

456

bromide to convert ammonium to nitrite. Using this method, an LOD of 1.9 µM was

457

obtained for ammonium with the linear range up to 57 µM. The method was validated by

458

determination of ammonium in several real seawater samples, with the results compared

459

with those obtained by standard batch methods, showing no significant differences.

460

Furthermore, applications of hypobromite oxidation spectrophotometry for the

21

461

simultaneous determination of inorganic nitrogen in soils and environmental waters have

462

also reported[53].

463 464

Analytical features of each spectrophotometric method which has been applied in seawater analysis from 1999 to 2019 are summarized in Table 1.

465

Table 1 Analytical features of spectrophotometric methods for ammonium determination

466

in seawater from 1999-2019

467 468

2.3 Fluorometric method

469

The fluorometric method involving reaction of ammonium with OPA is attractive due

470

to its high sensitivity. This method was first developed in 1971[58] for the determination

471

of amino acids, with amino acids reacting with OPA in the presence of 2-mercaptoethanol

472

(or borohydride) under alkaline conditions, to produce a fluorescent product. This method

473

rapidly became one of the most commonly applied methods for chromatographic analysis

474

of amino acids. Although the reaction was applied to ammonia analysis in 1974[45], the

475

reported use of this reaction has continued to focus on the determination of amino acids,

476

with few studies reporting the measurement of ammonium using this reaction in the last

477

15 years[45]. As the reaction can be used for the measurement of both ammonium and

478

amino acids, interference by amino acids should be considered during application for

479

ammonium analysis. Therefore, the gas diffusion method has been applied to separate

480

ammonium from interfering compounds, with the reaction first used for ammonium

481

analysis in seawater in 1991[59]. To improve the selectivity of this method, the reaction

482

was modified by replacing mercaptoethanol with sulfite, providing a method with better 22

483

sensitivity and selectivity for ammonium than for amino acids[60], with phosphate added

484

as buffer to adjust the pH of seawater samples. Since then, the reaction of ammonium,

485

OPA and sulfite has become increasingly popular and is the most widely reported method

486

for ammonium determination. Scheme of the chemical reaction of ammonium, OPA and

487

sulfite is shown in Fig. 4. A direct fluorometric method without the need for sample

488

separation was developed in 1997, for ammonium analysis in seawater and estuarine

489

waters using SFA and the reaction of ammonium, OPA and sulfite[61]. This method has

490

the benefit of being free from primary amine interference, with a low salinity interference

491

effect of less than 3% in the 0.2-35 salinity range. As this reaction requires 3-4 h to reach

492

equilibrium at ambient temperatures, a high reaction temperature of 75 Ⅱ was selected to

493

accelerate the reaction process. Different buffers which were compatible with seawater

494

were also tested as part of this study and the borate buffer was selected instead of

495

phosphate in the final study, as it is simple to prepare and provided adequate sensitivity.

496 497 498

Fig. 4 Scheme of the chemical reaction of ammonium-OPA-sulfite Holmes et al.[62] combined the reagents OPA, sulfite and borate to form a single

499

working reagent, and employed a manual fluorometric method with a large linear range

500

for ammonium analysis in seawater. The matrix effects and background fluorescence

501

were assessed and corrected without introducing substantial error. However, Taylor et

502

al.[63] corrected the equations reported by Holmes et al.[62], providing a new method for

503

measurement of matrix effects and background fluorescence. Li et al.[64] investigated the

504

use of fluorescence detection using a transversely illuminated liquid core waveguide 23

505

(LCW), coupled with FIA for ammonium fluorescence determination. Watson et al.[27]

506

modified this method using a GD unit to minimize the potential interference of the

507

seawater matrix, with application tested in Southern Ocean and Huon Estuary (Tasmania)

508

waters. Contamination by ammonia from laboratory and shipboard sources was avoided

509

by the use of special process, resulting in this method being more suitable for low level

510

ammonium measurements in open ocean. The LOD of this method was 7 nM, with linear

511

calibration up to the maximum tested level of 4 µM. Maruo et al.[65] combined

512

continuous flow analysis (CFA) with the OPA-sulfite reaction, with application for the

513

determination of ammonium in ice core and lake. The reaction of OPA and sulfite was

514

adapted to FIA by Aminot et al.[66] for in-situ application in fresh or saline natural waters,

515

providing an LOD of 30 nM with linear calibration across a wide range up to 100 µM.

516

Using this method, no adverse effect was observed with hydrogen carbonate, dissolved

517

oxygen or turbidity, while the interference of primary amines and the effect of salinity

518

over a range of 5-35 were not significant. A SIA system for rapid determination of

519

ammonium and phosphate in coastal waters was presented by Frank et al.[67], with a

520

sample throughput of 120 h-1. The system was applied on several cruises in the North Sea

521

[68], allowing nutrient mapping across coastal areas. Poulin & Pelletier[69] described a

522

microplate-based fluorescence fluorescence method for fast ammonium measurement in

523

different groups of samples, with a LOD of 50 nM found with reddish turbid water

524

samples from the south shore of the St. Lawrence Estuary. While this method showed

525

good adaptability in estuary waters, the high LOD makes this method inadequate for

526

application to open ocean samples. 24

527

Amornthammarong et al. performed a series of studies on ammonium measurement

528

using the fluorometric method. Firstly, Amornthammarong et al.[70] developed a hybrid

529

fluorometric flow analyzer for determination of ammonia in the atmosphere based on the

530

reaction of ammonium-OPA-sulfite. The method was then modified by combining

531

formaldehyde with the sulfite solution to achieve a higher sensitivity and more stable

532

reagent solution[7], which also reduced potential interference from species such as

533

amines and amino acids. Following this, a continuous flow shipboard analyzer was

534

developed for determination of ammonium in seawater at highly sensitivity (LOD of 1.1

535

nM), with negligible salinity effect and no refractive index effect. The sample throughput

536

was 3600 h-1 using a CFA system, which was used to examine the ammonium distribution

537

in Florida coastal waters surrounding a wastewater outfall point. However, it should be

538

noted that application of this method in low concentration ammonium environments

539

requires correction of interfering species, such as amino acids. Following then, an

540

autonomous batch analyzer (ABA) was included as a modification of the SIA system by

541

Amornthammarong et al.[71]. The addition of an ABA provided the advantage of a

542

mixing chamber, which overcomes the limitations of batch and continuous flow analyzers

543

by allowing higher sensitivity (LOD was 1 nM) due to complete and rapid mixing. Using

544

this method, calibration curves can be produced autonomously by auto-dilution of a

545

single stock standard solution. The ABA modified method was used to characterize

546

surface ammonium concentrations in the coastal waters of the Florida Keys and

547

southeastern coastal waters, with underway measurement in surface seawater from Key

548

West to the Boca Raton inlet (Florida, USA). The ABA system was later modified using 25

549

an optimized LED photodiode-based fluorescence detector[72]. This modification

550

resulted in greater sensitivity and a considerably smaller equipment size than previous

551

systems, while incorporating a pre-filtering component, enabling measurements in turbid,

552

sediment-laden waters with an LOD of 10 nM. The portable analyzer was tested at three

553

shallow South Florida sites, for analysis of diurnal cycles and the potential transport of

554

ammonium into coastal waters. Bey et al.[73] described a high resolution fluorometric

555

system for ammonium measurement in oligotrophic seawater, which was applied to

556

analysis of low ammonium concentration seawater in the North Atlantic Ocean. The

557

effects of salinity, amines, amino acids and potential interference from particles or algae,

558

were investigated. Results showed that the method was sensitive to salinity variations

559

especially in low concentration ammonium environments, reducing the signal by up to

560

85% at 5 nM, with high amino acid or amine concentrations also affecting ammonium

561

measurements. In addition, phytoplankton blooms were found to have a significant

562

impact, altering the ammonium signal by up to 12%. Horstkotte et al.[4] designed a

563

portable multi-pumping flow analyzer using an rFIA concept for shipboard monitoring.

564

The system was found to be reliable and robust across a wide range of pH, salinity and

565

temperature conditions, as well as gas concentrations, with the sensitivity improved by

566

the use of a photomultiplier tube (PMT). In order to further improve the sensitivity of the

567

method, Zhu et al.[74] developed a flow-batch system combining SPE with fluorescence

568

detection for ultra-trace ammonium concentration (<1 nM) measurements in seawater.

569

The fluorescent product was extracted using an SPE cartridge (HLB), separating the

570

analyte complex from the seawater matrix and enriching ammonium, with the extracted 26

571

compounds then rapidly with ethanol and measured by fluorometry. This method

572

provided a high sensitivity with an LOD of 0.7 nM and 1.2 nM in land-based and

573

shipboard laboratory analysis, respectively, without salinity and matrix interference. This

574

method was applied to the generation of a high-resolution vertical profile of ammonium

575

in the South China Sea, along with the distribution of ammonium in the surface seawaters

576

in the region. In a recent study by Zhu et al.[75] a home-made portable fluorescence

577

detector was described, comprised of a UV-LED, two band pass filters, a PMT, a

578

modified flow cell and an electronic circuit with a constant voltage and current supply.

579

The custom-made detector could efficiently avoid interference to the signal from air

580

bubbles and provided improved sensitivity compared to the commercially available

581

system one. Combining the FIA technique with an underway sampling system, the

582

detector was applied to underway analysis of the distribution of ammonium in surface

583

waters of the Jiulongjiang Estuary

584

Hu et al.[76] described a manual operation method based on the reaction of

585

ammonium-OPA-sulfite, with the reagent EDTA-NaOH used as a buffer instead of

586

sodium tetraborate. This method provided high sensitivity with an LOD of 9.9 nM and

587

was applied to the measurement of ammonium in natural river water samples. A novel

588

fluorescent reagent, 4-methoxyphthalaldehyde (MOPA) was reported by Liang et al.[77],

589

which was developed by adding an electron-donating methoxy group to the benzene ring

590

of OPA. MOPA was applied in the reaction of ammonium and sulfite as a substitute for

591

OPA and was found to rapidly react at room temperature. A novel fluorescence method

592

was then developed using MOPA for ammonium measurement in river water and 27

593

seawater matrices, with an LOD of 5.8 nM obtained, showing an increase in sensitivity

594

compared to the OPA method. Later, another novel fluorescent reagent,

595

4,5-dimethoxyphthalaldehyde (M2OPA), was synthesized by adding two methoxy groups

596

to the OPA benzene ring, as reported by Zhang et al.[78]. Using a custom-made hand-held

597

portable fluorometer with a laser diode light source, a novel method was developed for

598

sensitively detecting ammonium in river water and seawater, with no additional reaction

599

heating required for direct field analysis.

600

Some other methods have also been combined, such as matrix separation techniques

601

with fluorometric methods, which have been applied in ammonium measurement and will

602

be discussed in the following section. Analytical features of each fluorescence method

603

which has been applied in seawater analysis from 1999 to 2019 are summarized in Table

604

2.

605

Table 2 Analytical features of fluorescence methods for ammonium determination in

606

seawater from 1999-2019

607 608

2.4 Methods based on matrix separation

609

The complexity of the seawater matrix may cause significant interference during the

610

measurement of ammonium in seawater. Matrix effects include pH, buffering capacity,

611

ionic strength, salinity effects, interference from naturally occurring nitrogen-containing

612

compounds, turbidity and other interferences from particles or algae. These effects are a

613

major limitation in the application of most methods for ammonium analysis in seawater

614

environments. However, if ammonium can be separated from the sample matrix, then 28

615

matrix effects can be minimized or removed, allowing accurate measurement of

616

ammonium concentrations in seawater matrices. A key advantage of matrix separation

617

methods is that ammonium in samples is concentrated to improve the sensitivity of

618

analysis. Therefore, several approaches such as gas diffusion (GD) and purge-and-trap

619

(P&T) systems have been developed and applied in the determination of ammonium in

620

seawater matrices.

621

2.4.1 Gas diffusion

622

Of the commonly used membrane-based separation techniques (dialysis, GD and

623

pervaporation) GD has been frequently applied in combination with flow analysis

624

techniques in environmental analysis, due to its simplicity, selectivity, high-enrichment

625

factor potential and low use of solvents. GD has commonly been used to separate

626

ammonium from seawater samples prior to detection, following a similar mechanism as

627

described for the ammonia gas sensing probe in section 2.1. Under alkaline conditions,

628

ammonium in the sample is converted into ammonia in a donor stream, which then

629

diffuses across a hydrophobic gas permeable membrane into an acceptor stream and is

630

absorbed into a receiving solution, which can be measured by various methods.

631

The GD unit has been widely used in combination with flow analysis and was first

632

applied to the determination of ammonia by Růžička & Hansen[79]. Several studies have

633

applied GD coupled FIA analysis for ammonium measurement in seawater. Watson et

634

al.[27] developed a FIA system coupled with GD and fluorometry for ammonium

635

determination in seawater, with the method applied in the Southern Ocean and Huon

636

Estuary (Tasmania, Australia). As ammonium in samples is converted into ammonia and 29

637

diffuses over the membrane, a pH change is induced and subsequently a color change

638

occurs in the acceptor stream. The ammonia absorbed in the acceptor stream can then be

639

determined using a pH indicator reagent (bromothymol blue solution) by

640

spectrophotometry. Based on this reaction, Gray et al.[80] developed a hybrid reagent

641

injection flow analysis system (same as rFIA) where the reagent sodium hydroxide was

642

injected into a flowing sample stream to reduce reagent consumption and waste

643

production. The interference of dissolved carbon dioxide in samples was minimized, even

644

in the presence of a wide alkalinity range (28.8-131 mg CaCO3 L-1), by online adjustment

645

of the sample pH. Using the same detection method, SIA has been employed as an

646

alternative to rFIA, in a novel method developed for ammonium determination in

647

transitional and coastal waters, combining GD and spectrophotometry to measure the

648

change in absorbance of the pH indicator[81]. As an alternative to the FIA/SIA system,

649

Oliveira et al.[82] proposed the use of a multi-commuted FIA system with a

650

multi-channel propulsion unit added prior to detection, for ammonium determination in

651

surface waters and tap water. The use of this system generates a positive pressure to

652

facilitate mass separation by GD and avoid the formation of air bubbles. Following this,

653

the method was modified for application in seawater and estuarine waters under a wide

654

range of salinity conditions[83]. Several complexing agents were evaluated for their

655

ability to prevent precipitation of metallic hydroxides and an extensive study of possible

656

interfering species was performed. An LOD of 1.3 µM was obtained using this method,

657

which may be inadequate for application in unpolluted seawater and open water

658

environments. The multi-commuted (or multi-syringe) FIA system has also been coupled 30

659

with a conductivity detector for ammonium determination in coastal waters [84, 85]. Plant

660

et al.[86] demonstrated the use of an in-situ analyzer combined with a GD cell and

661

conductivity detector. Micro-solenoid pumps were used to propel the sample and reagents

662

through the system, using a similar process as described for the multi-pumping flow

663

system. The advantages of this system are high sensitivity (LOD was 10 nM) and stability

664

(at least 30 d), with suitability for fixed location monitoring of ammonium in estuarine

665

and coastal waters at depths of up to 3 m. This system has been successfully deployed on

666

coastal moorings, in benthic flux chambers and on a drifter 500 km west of Monterey Bay

667

(California, US). In order to improve the sensitivity of GD based methods, Kodama et

668

al.[34] developed a continuous flow method coupled with GD, LWCC and IPB

669

spectrophotometric methods for low concentration ammonium measurement in seawater.

670

An LOD of 5.5±1.8 nM was obtained, making this method more suitable for the

671

determination of horizontal distributions and spatial variations of ammonium in

672

oligotrophic oceans. Recently, Šraj et al.[87] developed a simple environmentally-

673

friendly ammonium analyzer (SEA) based on programmable flow and GD

674

spectrophotometric methods using the pH indicator bromothymol blue for trace level

675

ammonium determination in marine waters. The degree of pre-concentration of

676

ammonium in the acceptor stream was determined and the SEA analyzer utilized

677

enhanced flow manipulation, allowing the sensitivity of the method to be tailored to the

678

required concentration range via simple adjustment of the sample volume. In addition, the

679

wide linear calibration range makes this method versatile and potentially applicable for

680

use in coastal and oceanic environments. Furthermore, a novel passive sampler based on 31

681

GD was also described by Šraj et al.[87], providing a time-weighted average

682

concentration of dissolved molecular ammonia in marine waters over a period of 3 days.

683

Good agreement was found between passive and spot sampling results in both cases,

684

indicating this method may be applicable for long-term ammonia monitoring in estuarine

685

and coastal waters.

686

2.4.2 Purge-and-trap

687

P&T is another well-established separation technique which is often used for the

688

determination of volatile organic compounds especially in wastewater and other complex

689

water matrices. The mechanism of P&T involves passing inert gas bubbles through the

690

water sample to force purgeable compounds from the aqueous phase to the vapor phase,

691

which are then caught on a trap for further analysis[88]. For ammonium measurement in

692

seawater, it is also necessary to add sodium hydroxide to the water sample to convert

693

ammonium into gaseous ammonia (in a similar manner to GD), with the P&T system

694

trapped ammonium then potentially determined via various methods. Compared with GD,

695

the P&T system can process large sample volumes in the pretreatment progress,

696

increasing the enrichment factor and therefore, improving the sensitivity of the method.

697

Wang et al.[88] developed a novel P&T pre-concentrator for trace ammonium

698

determination in high-salinity water by ion chromatography (IC). Under optimum

699

conditions an LOD of 75 nM was obtained, which could be lowered to 7.5 nM) using a

700

larger sample loop volume. This method was applied to natural seawater samples

701

collected from coastal areas. This method was modified by Lin et al.[89] , resulting in a

702

significantly enhanced performance and efficiency for the quantification of ammonium, a 32

703

reduced background signal and an increase in the maximum overall collection efficiency

704

from 64% to 97%. This method could be applied to the measurement of ammonium over

705

a wide dynamic range (5 orders of magnitude) of concentrations from 50 nM to >10 mM.

706

In addition, interference from amino acids was assessed and with the exception of

707

asparagine and glutamine, minor interference was observed with 20 native amino acids

708

although no significant effects were reported overall. Ferreira et al.[90] developed an

709

ultrasound-assisted P&T extraction system with IC for the simultaneous determination of

710

amines and ammonium in high salinity waters, although the LOD of this method was high

711

at almost 14 µM. An improved method was proposed by Zhu et al.[91] using a

712

home-made P&T pretreatment system coupled with FIA and fluorometry. Using this

713

method, the influence of primary amines and amino acids were found to be negligible,

714

with an LOD of 7.4 nM obtained and a linear calibration range of 10-400 nM, although

715

deionized water was used for standard preparation instead of matrix-matched standards.

716

2.4.3 Other separation approaches

717

There are still some other separation approaches (such as ion-exchange

718

chromatography or pervaporation) have been applied to ammonium measurement in

719

environmental waters, although these have rarely included seawater analysis. An

720

improved high-performance liquid chromatographic (HPLC) method was developed for

721

ammonium and primary amine measurement in seawater, using a high-efficiency

722

cation-exchange column combined with modified buffer and reagent solutions[92]. IC

723

using a high-capacity cation-exchange column has also been used to separate ammonium

724

from high concentration ratios of sodium to ammonium[93]. To remove amino acid 33

725

interference, a post-column derivatization method based on the reaction of ammonium,

726

OPA and sulfite has been used, providing an LOD of 100 nM. Ferreira et al.[94] used

727

steam distillation as a separation technique and developed a novel method using IC for

728

the determination of ammonium, monomethylamine and monoethylamine in saline waters.

729

The uses of SPE or solvent extraction are commonly used separation approaches. A

730

method involving headspace single-drop microextraction (SDME) and capillary

731

electrophoresis (CE) was developed by Pranaitytė et al.[95] for ammonium determination

732

in environmental and biological samples. This method resulted in a 14-fold level of

733

enrichment in 20 min, providing an LOD of 1.5 µM with a linear calibration range of

734

5-100 µM. Muniraj et al.[96] developed an automated headspace dynamic in-syringe

735

liquid phase microextraction (LPME) technique using in-situ derivatization coupled with

736

liquid chromatography fluorescence detection. This method was applied to the

737

determination of ammonium in lake water, river water and tap water, providing an LOD

738

of 330 nM. A novel system using an automated syringe technique was developed by

739

Šrámková et al.[97] based on headspace SDME, which was used to determine ammonium

740

according to the color change of an acid-base-indicator drop.

741 742

Analytical features of methods based on matrix separation for ammonium analysis in seawater from 1999 to 2019 are summarized in Table 3.

743

Table 3 Analytical features of methods based on matrix separation for ammonium

744

determination in seawater from 1999-2019

745

3. Ammonium-free seawater preparation method

34

746

One of the limitations to the quantitative estimation of low concentration ammonium is

747

the availability of ammonium-free seawater (AFSW). For most of the proposed methods

748

for ammonium measurement in seawater, pure water cannot be used to prepare standard

749

solutions. As seawater and pure water exhibit differences in salinity, refractive index,

750

ionic strength, pH and buffering capacity, both the standard solutions and washing/carrier

751

solutions used in the flow-based methods should be prepared in AFSW to minimize

752

matrix effects and reduce the background signal of the matrix[98]. LNSW collected from

753

the surface of oligotrophic oceans are typically used as a substitute for AFSW and has

754

been applied in many studies. Li et al.[32] described a method to prepare low ammonium

755

seawater (LASW), in which NaOH was added to LNSW collected from the surface of the

756

Gulf Stream, until a small amount of precipitate was formed. Following this, the seawater

757

mixture was continually agitated and heated to 60Ⅱ to evaporate the gaseous ammonia.

758

The solution was sealed and cooled to room temperature, before being filtered through a

759

0.45 µm membrane filter to form LASW. However, this method destroys the seawater

760

matrix due to a large change in pH, salinity and carbonate buffering capacity, making the

761

prepared LASW ineffective. Clark et al.[30] developed another LASW preparation

762

method based on the diffusion of ammonia through a PTFE membrane into acid. An

763

LNSW sample (2 L) collected from an oligotrophic ocean region (ammonium

764

concentrations ranging from 10-100 nM) was filtered through a 0.2 µm membrane filter

765

and placed in a borosilicate reservoir vessel, with continual agitation using a magnetic

766

stirrer. The vessel was sealed with a gas tight cap fitted with a 3-valve adaptor and the

767

sample was pumped from the reservoir through 8×1 m lengths of PTFE tubing and 35

768

immersed in 10% (v/v) HCl. The treated seawater was returned to the vessel and

769

recirculated through this system for >24 h to obtain AFSW. The pH of the prepared

770

LASW slightly decreased (typically by 0.1-0.5 pH units), maintaining the original

771

seawater matrix. Almost all ammonium was converted to ammonia when the sample pH

772

was above 11.5[96] (ammonium ion pKa 9.24). However, trace ammonia cannot be easily

773

and rapidly purged from water even under high pH condition (pH 13)[88], due to the high

774

solubility of ammonia in water. Zhu et al.[98] described a novel AFSW preparation

775

method, using a preparation system consisting of only a peristaltic pump and an HLB

776

cartridge. The original seawater sample was collected in a HDPE bottle, with OPA and

777

sulfite added and the mixture left to react for more than 16 h to complete the reaction

778

between ammonium, OPA and sulfite. The seawater mixture was then passed through the

779

HLB cartridge via a peristaltic pump, while the ammonium, OPA and sulfite reaction

780

product was retained by the HLB cartridge and the eluent was considered to be AFSW.

781

The eluting efficiency of the HLB cartridge and the ammonium removal efficiency under

782

different reagent conditions were assessed. Results showed that the ammonium

783

concentration in the prepared AFSW was much lower than in surface seawater collected

784

from the South China Sea.

785

4. Conclusions

786

In order to better understand the development of typically used methodologies in

787

seawater analysis and help select the most appropriate method to meet specific

788

requirements, this review summarizes the development of the most commonly used

789

methodologies for ammonium determination in seawater, including ion-selective 36

790

electrode methods, spectrophotometric methods, fluorometric methods and methods

791

based on matrix separation.

792

The ISE method is most suitable for rapid monitoring applications, early warning

793

monitoring and in-situ direct analysis of ammonium in aquaculture samples or polluted

794

natural water systems, as unlike other methods it does not require dilution of samples

795

with relatively high concentrations of ammonium, due to the relatively low upper limit.

796

However, the ISE method is rarely applied to determination of low ammonium

797

concentrations in seawater as most ISE sensors are limited by interference from other ions,

798

low sensitivity, inadequate long-term stability, unstable performance and poor

799

reproducibility. Although great progress has been made to address these limitations, much

800

work is still needed to improve the sensor performance for practical application and

801

commercialization.

802

Spectrophotometric methods are commonly used for ammonium determination and

803

have been adopted as standard methods. Dependency on toxic reagents and low

804

sensitivity are the main drawbacks of the traditional IPB method using phenol. However,

805

these problems have been solved with method optimization and modification. The use of

806

phenol, a toxic reagent, can be mitigated by reducing the usage volume of phenol or

807

avoided by using an alternative reagent such as OPP. In addition, sensitivity can be

808

improved by using flow analysis techniques and by applying LWCC or preconcentration

809

techniques, such as SPE. The basic spectrophotometry methods using phenol, or the IPB

810

method using OPP and hypobromite oxidation, seem to be most suitable for application to

811

seawater analysis. 37

812

Fluorometric methods are dominantly used for ammonium analysis due to their high

813

sensitivity, without the need for enrichment techniques. When combined with flow

814

analysis techniques, fluorometric methods are not only used in laboratory analysis, but

815

can also be applied in shipboard or in-situ measurements, both in coastal and open ocean

816

waters. When the fluorometric method is applied in combination with a continuous flow

817

analysis system, the advantages of high sensitivity and high sample throughput make this

818

method more suitable for high resolution underway analysis of ammonium in surface

819

seawater. The newly developed fluorescent reagents MOPA and M2OPA still require

820

further investigation to confirm their long-term stability, as well as further application in

821

natural seawater environments.

822

The fluorometric method coupled with matrix separation techniques can minimize or

823

remove interference caused by seawater matrices, which is useful for accurate

824

measurement of ammonium concentrations especially in complex seawater samples. The

825

P&T system requires an inert gas and a complex processing unit, which can be used in

826

laboratory settings but may not be suitable for field-based applications. Compared with

827

the P&T system, the GD system has better prospects for practical application, in

828

laboratory, on-line or shipboard application environments. However, the mass transfer

829

efficiency of the GD unit is debatable and the sensitivity of the method is not high enough

830

for low concentration monitoring in open ocean environments. Therefore, methods based

831

on matrix separation are more suitable for ammonium determination in coastal and

832

estuarine waters.

833

Acknowledgments 38

834

This work was supported by the National Natural Science Foundation of China (Grant

835

number 41606101) and Zhejiang Provincial Natural Science Foundation of China (Grant

836

number LGC19D060002).

837

References

838 839 840 841 842 843 844 845 846 847 848 849 850 851 852 853 854 855 856 857 858 859 860 861 862 863 864 865 866 867 868 869 870 871 872

[1] F.J. Millero, Chemical oceanography, CRC press, (2016). [2] M. Whitfield, The hydrolysis of ammonium ions in sea water-experimental confirmation of predicted constants at one atmosphere pressure. J. Mar. Biol. Assoc. UK 58 (1978) pp. 781-786. [3] R.W. Eppley, B.J. Peterson, Particulate organic matter flux and planktonic new production in the deep ocean, Nature 282 (1979) pp. 677. [4] B. Horstkotte, C.M. Duarte, V. Cerda, A miniature and field-applicable multipumping flow analyzer for ammonium monitoring in seawater with fluorescence detection, Talanta 85 (2011) pp. 380-385. [5] C. Molins-Legua, S. Meseguer-Lloret, Y. Moliner-Martinez, P. Campíns-Falcó, A guide for selecting the most appropriate method for ammonium determination in water analysis, TrAC, Trends Anal. Chem. 25 (2006) pp. 282-290. [6] L.O.C. Šraj, M.I.G. Almeida, S.E. Swearer, S.D. Kolev, I.D. McKelvie, Analytical challenges and advantages of using flow-based methodologies for ammonia determination in estuarine and marine waters, TrAC, Trends Anal. Chem. 59 (2014) pp. 83-92. [7] N. Amornthammarong, J.Z. Zhang, Shipboard fluorometric flow analyzer for high-resolution underway measurement of ammonium in seawater, Anal. Chem. 80 (2008) pp. 1019-1026. [8] H.J. Liew, A.K. Sinha, C.M. Nawata, R. Blust, C.M. Wood, G. De Boeck, Differential responses in ammonia excretion, sodium fluxes and gill permeability explain different sensitivities to acute high environmental ammonia in three freshwater teleosts, Aquat. Toxicol. 126 (2013) pp. 63-76. [9] P.J. Statham, Nutrients in estuaries - An overview and the potential impacts of climate change, Sci. Total Environ. 434 (2012) pp. 213-227. [10] J. Ma, L. Adornato, R.H. Byrne, D. Yuan, Determination of nanomolar levels of nutrients in seawater, TrAC, Trends Anal. Chem. 60 (2014) pp. 1-15. [11] J. Ma, D. Yuan, K. Lin, S. Feng, T. Zhou, Q. Li, Applications of flow techniques in seawater analysis: A review, Trends Environ. Anal. Chem. 10 (2016) pp. 1-10. [12] L.M. Nollet, L.S. De Gelder, Handbook of water analysis, CRC press, (2014). [13] G.A. Crespo, Recent advances in ion-selective membrane electrodes for in situ environmental water analysis, Electrochim. Acta 245 (2017) pp. 1023-1034. [14] L. Zhou, C.E. Boyd, Comparison of Nessler, phenate, salicylate and ion selective electrode procedures for determination of total ammonia nitrogen in aquaculture, Aquaculture 450 (2016) pp. 187-193. [15] X. Chen, G. Zhou, S. Mao, J. Chen, Rapid detection of nutrients with electronic sensors: a review, Environ. Sci.: Nano 5 (2018) pp. 837-862. [16] T.R. Gilbert, A.M. Clay, Determination of ammonia in aquaria and in sea water using the ammonia electrode, Anal. Chem. 45 (1973) pp. 1757-1759. 39

873 874 875 876 877 878 879 880 881 882 883 884 885 886 887 888 889 890 891 892 893 894 895 896 897 898 899 900 901 902 903 904 905 906 907 908 909 910 911 912 913 914 915

[17] A. Merks, Determination of ammonia in sea water with an ion-selective electrode, Neth. J. Sea Res. 9 (1975) pp. 371-375. [18] E. Moschou, U.A. Lasarte, M. Fouskaki, N. Chaniotakis, N. Papandroulakis, P. Divanach, Direct electrochemical flow analysis system for simultaneous monitoring of total ammonia and nitrite in seawater, Aquacult. Eng. 22 (2000) pp. 255-268. [19] Y. Wen, Y. Mao, Z. Kang, Q. Luo, Application of an ammonium ion-selective electrode for the real-time measurement of ammonia nitrogen based on pH and temperature compensation, Measurement 137 (2019) pp. 98-101. [20] C.W. Jang, Y.T. Byun, Y.M. Jhon, Detection of 10 nM ammonium ions in 35‰ NaCl solution by carbon nanotube based sensors, J. Nanosci. Nanotechnol. 12 (2012) pp. 1765-1769. [21] L. Ding, J. Ding, B. Ding, W. Qin, Solid-contact potentiometric sensor for the determination of total ammonia nitrogen in seawater, Int. J. Electrochem. Sci. 12 (2017) pp. 3296-3308. [22] M. Takahashi, K. Nakamura, J. Jin, Study on the indirect electrochemical detection of ammonium ion with in situ electrogenerated hypobromous acid, Electroanalysis 20 (2008) pp. 2205-2211. [23] P.L. Searle, The Berthelot or indophenol reaction and its use in the analytical chemistry of nitrogen. A review, Analyst 109 (1984) pp. 549-568. [24] M.P.E. Berthelot, Violet d'aniline, Rep. Chim. Appl. 1 (1859) p. 284. [25] T. Shoji, E. Nakamura, Collection of indonaphthol blue on a membrane filter for the spectrophotometric determination of ammonia with 1-naphthol and dichloroisocyanurate, Anal. Sci. 26 (2010) pp. 779-783. [26] A. Aminot, D. Kirkwood, R. Kerouel, Determination of ammonia in seawater by the indophenol-blue method: Evaluation of the ICES NUTS I/C 5 questionnaire, Mar. Chem. 56 (1997) pp. 59-75. [27] R.J. Watson, E.C.V. Butler, L.A. Clementson, K.M. Berry, Flow-injection analysis with fluorescence detection for the determination of trace levels of ammonium in seawater, J. Environ. Monit. 7 (2005) pp. 37-42. [28] S.C. Pai, Y.J. Tsau, T.I. Yang, pH and buffering capacity problems involved in the determination of ammonia in saline water using the indophenol blue spectrophotometric method, Anal. Chim. Acta 434 (2001) pp. 209-216. [29] M.A. Brzezinski, Colorimetric determination of nanomolar concentrations of ammonium in seawater using solvent extraction, Mar. Chem. 20 (1987) pp. 277-288. [30] D.R. Clark, T.W. Fileman, I. Joint, Determination of ammonium regeneration rates in the oligotrophic ocean by gas chromatography/mass spectrometry, Mar. Chem. 98 (2006) pp. 121-130. [31] G. Chen, M. Zhang, Z. Zhang, Y. Huang, D. Yuan, On-line solid phase extraction and spectrophotometric detection with flow technique for the determination of nanomolar level ammonium in seawater samples, Anal. Lett. 44 (2011) pp. 310-326. [32] Q.P. Li, J.Z. Zhang, F.J. Millero, D.A. Hansell, Continuous colorimetric determination of trace ammonium in seawater with a long-path liquid waveguide capillary cell, Mar. Chem. 96 (2005) pp. 73-85. [33] Y. Zhu, D. Yuan, Y. Huang, J. Ma, S. Feng, K. Lin, A modified method for on-line determination of trace ammonium in seawater with a long-path liquid waveguide capillary cell and spectrophotometric detection, Mar. Chem. 162 (2014) pp. 114-121. 40

916 917 918 919 920 921 922 923 924 925 926 927 928 929 930 931 932 933 934 935 936 937 938 939 940 941 942 943 944 945 946 947 948 949 950 951 952 953 954 955 956

[34] T. Kodama, T. Ichikawa, K. Hidaka, K. Furuya, A highly sensitive and large concentration range colorimetric continuous flow analysis for ammonium concentration, J. Oceanogr. 71 (2015) pp. 65-75. [35] F. Hashihama, J. Kanda, A. Tauchi, T. Kodama, H. Saito, K. Furuya, Liquid waveguide spectrophotometric measurement of nanomolar ammonium in seawater based on the indophenol reaction with o-phenylphenol (OPP), Talanta 143 (2015) pp. 374-380. [36] J. Ma, P. Li, K. Lin, Z. Chen, N. Chen, K. Liao, D. Yuan, Optimization of a salinity-interference-free indophenol method for the determination of ammonium in natural waters using o-phenylphenol, Talanta 179 (2018) pp. 608-614. [37] A. Kempers, C. Kok, Re-examination of the determination of ammonium as the indophenol blue complex using salicylate, Anal. Chim. Acta 221 (1989) pp. 147-155. [38] H. Muraki, K. Higuchi, M. Sasaki, T. Korenaga, K. Tôei, Fully automated system for the continuous monitoring of ammonium ion in fish farming plant sea water by flow-injection analysis, Anal. Chim. Acta 261 (1992) pp. 345-349. [39] F. Jüttner, Interference with ammonium determination by the indophenol-type reaction of salicylate and dichloroisocyanurate, Fresenius J. Anal. Chem. 363 (1999) pp. 128-129. [40] P.T.T. Le, C. Boyd, Comparison of phenate and salicylate methods for determination of total ammonia nitrogen in freshwater and saline water, J. World Aquacult. Soc. 43 (2012) pp. 885-889. [41] J. Kanda, Determination of ammonium in seawater based on the indophenol reaction with o-phenylphenol (OPP), Water Res. 29 (1995) pp. 2746-2750. [42] K. Lin, P. Li, Q. Wu, S. Feng, J. Ma, D. Yuan, Automated determination of ammonium in natural waters with reverse flow injection analysis based on the indophenol blue method with o-phenylphenol, Microchem. J. 138 (2018) pp. 519-525. [43] J. Ma, P. Li, Z. Chen, K. Lin, N. Chen, Y. Jiang, J. Chen, B. Huang, D. Yuan, Development of an integrated syringe-pump-based environmental-water analyzer (iSEA) and application of it for fully automated real-time determination of ammonium in fresh water, Anal. Chem. 90 (2018) pp. 6431-6435. [44] P. Li, Y. Deng, H. Shu, K. Lin, N. Chen, Y. Jiang, J. Chen, D. Yuan, J. Ma, High-frequency underway analysis of ammonium in coastal waters using an integrated syringe-pump-based environmental-water analyzer (iSEA), Talanta 195 (2019) pp. 638-646. [45] S.S. Goyal, D.W. Rains, R.C. Huffaker, Determination of ammonium ion by fluorometry or spectrophotometry after on-line derivatization with o-phthalaldehyde, Anal. Chem. 60 (1988) pp. 175-179. [46] Y. Liang, C. Yan, Q. Guo, J. Xu, H. Hu, Spectrophotometric determination of ammonia nitrogen in water by flow injection analysis based on NH3-o-phthalaldehyde-Na2SO3 reaction, Anal. Chem. Res. 10 (2016) pp. 1-8. [47] F.A. Richards, R.A. Kletsch, The spectrophotometric determination of ammonia and labile amino compounds in fresh and seawater by oxidation to nitrite, Techinical report No. 97, University of Washington press, (1964). [48] K. Matsunaga, M. Nishimura, A rapid and sensitive method for determination of submicrogram amounts of ammonia in fresh and sea waters, Anal. Chim. Acta 73 (1974) pp. 204-208.

41

957 958 959 960 961 962 963 964 965 966 967 968 969 970 971 972 973 974 975 976 977 978 979 980 981 982 983 984 985 986 987 988 989 990 991 992 993 994 995 996 997 998 999

[49] R. Liu, H. Wang, A. Sun, D. Liu, Reversed flow injection spectrophotometric determination of trace amount of ammonia in natural water by oxidation of ammonia to nitrite, Talanta 45 (1997) pp. 405-410. [50] Y. Minamiya, I. Nukatsuka, K. Ohzeki, Determination of ammonia by solid-phase spectrophotometry, Bunseki Kagaku 50 (2001) pp. 341-344. [51] B. Sun, R. John, H. Zhao, Development of a fully automated inorganic nitrogen analyzer for continuous, unattended monitoring of water quality, Lab. Rob. Autom. 12 (2000) pp. 312-316. [52] A. Tovar, C. Moreno, M.P. Mánuel-Vez, M. Garcı́a-Vargas, A simple automated method for the speciation of dissolved inorganic nitrogen in seawater, Anal. Chim. Acta 469 (2002) pp. 235-242. [53] C.E.L. Pasquali, P.F. Hernando, J.D. Alegria, Spectrophotometric simultaneous determination of nitrite, nitrate and ammonium in soils by flow injection analysis, Anal. Chim. Acta 600 (2007) pp. 177-182. [54] S. Ringuet, L. Sassano, Z.I. Johnson, A suite of microplate reader-based colorimetric methods to quantify ammonium, nitrate, orthophosphate and silicate concentrations for aquatic nutrient monitoring, J. Environ. Monit. 13 (2011) pp. 370-376. [55] F. Azzaro, M. Galletta, Automatic colorimetric analyzer prototype for high frequency measurement of nutrients in seawater, Mar. Chem. 99 (2006) pp. 191-198. [56] B. Bucur, M. Catala Icardo, J. Martinez Calatayud, Spectrophotometric determination of ammonium by an rFIA assembly, Rev. Roum. Chim. 51 (2006) p. 101. [57] T. Shoji, E. Nakamura, Flow injection analysis with spectrophotometry for ammonium ion with 1-naphthol and dichloroisocyanurate, J. Flow Injection Anal. 26 (2009) p. 37. [58] M. Roth, Fluorescence reaction for amino acids, Anal. Chem. 43 (1971) pp. 880-882. [59] R.D. Jones, An improved fluorescence method for the determination of nanomolar concentrations for ammonium in natural waters, Limnol. Oceanogr. 36 (1991) pp. 814-819. [60] G.F. Zhang, P.K. Dasgupta, Fluorometric measurement of aqueous ammonium ion in a flow injection system, Anal. Chem. 61 (1989) pp. 408-412. [61] R. Kérouel, A. Aminot, Fluorometric determination of ammonia in sea and estuarine waters by direct segmented flow analysis, Mar. Chem. 57 (1997) pp. 265-275. [62] R.M. Holmes, A. Aminot, R. Kérouel, B.A. Hooker, B.J. Peterson, A simple and precise method for measuring ammonium in marine and freshwater ecosystems, Can. J. Fish. Aquat. Sci. 56 (1999) pp. 1801-1808. [63] B.W. Taylor, C.F. Keep, R.O. Hall, B.J. Koch, L.M. Tronstad, A.S. Flecker, A.J. Ulseth, Improving the fluorometric ammonium method: matrix effects, background fluorescence, and standard additions, J. N. Am. Benthol. Soc. 26 (2007) pp. 167-177. [64] J. Li, P.K. Dasgupta, G.F. Zhang, Transversely illuminated liquid core waveguide based fluorescence detection: Fluorometric flow injection determination of aqueous ammonium/ammonia, Talanta 50 (1999) pp. 617-623. [65] M. Maruo, E. Nakayama, H. Obata, K. Kamiyama, T. Kimoto, Application of the flow-through analyses of ammonia and calcium in ice core and fresh water by fluorometric detection, Field Anal. Chem. Tech. 5 (2001) pp. 29-36. [66] A. Aminot, R. Kérouel, D. Birot, A flow injection-fluorometric method for the determination of ammonium in fresh and saline waters with a view to in situ analyses, Water Res. 35 (2001) pp. 1777-1785. 42

1000 1001 1002 1003 1004 1005 1006 1007 1008 1009 1010 1011 1012 1013 1014 1015 1016 1017 1018 1019 1020 1021 1022 1023 1024 1025 1026 1027 1028 1029 1030 1031 1032 1033 1034 1035 1036 1037 1038 1039 1040 1041 1042

[67] C. Frank, F. Schroeder, R. Ebinghaus, W. Ruck, A fast sequential injection analysis system for the simultaneous determination of ammonia and phosphate, Microchim. Acta 154 (2006) pp. 31-38. [68] C. Frank, F. Schroeder, Using sequential injection analysis to improve system and data reliability of online methods: determination of ammonium and phosphate in coastal waters, J. Anal. Methods Chem. 2007 (2007) Article ID 49535. [69] P. Poulin, E. Pelletier, Determination of ammonium using a microplate-based fluorometric technique, Talanta 71 (2007) pp. 1500-1506. [70] N. Amornthammarong, J. Jakmunee, J. Li, P.K. Dasgupta, Hybrid fluorometric flow analyzer for ammonia, Anal. Chem. 78 (2006) pp. 1890-1896. [71] N. Amornthammarong, J.Z. Zhang, P.B. Ortner, An autonomous batch analyzer for the determination of trace ammonium in natural waters using fluorometric detection, Anal. Methods 3 (2011) pp. 1501-1506. [72] N. Amornthammarong, J.Z. Zhang, P.B. Ortner, J. Stamates, M. Shoemaker, M.W. Kindel, A portable analyser for the measurement of ammonium in marine waters, Environ. Sci.: Proc. Imp. 15 (2013) pp. 579-584. [73] S.K.A.K. Bey, D.P. Connelly, F.E. Legiret, A.J.K. Harris, M.C. Mowlem, A high-resolution analyser for the measurement of ammonium in oligotrophic seawater, Ocean Dynam. 61 (2011) pp. 1555-1565. [74] Y. Zhu, D. Yuan, Y. Huang, J. Ma, S. Feng, A sensitive flow-batch system for on board determination of ultra-trace ammonium in seawater: Method development and shipboard application, Anal. Chim. Acta 794 (2013) pp. 47-54. [75] Y. Zhu, J. Chen, X. Shi, D. Yuan, S. Feng, T. Zhou, Y. Huang, Development and application of a portable fluorescence detector for shipboard analysis of ammonium in estuarine and coastal waters, Anal. Methods 10 (2018) pp. 1781-1787. [76] H. Hu, Y. Liang, S. Li, Q. Guo, C. Wu, A modified o-phthalaldehyde fluorometric analytical method for ultra-trace ammonium in natural waters using EDTA-NaOH as buffer, J. Anal. Methods Chem. 2014 (2014) Article ID 728068. [77] Y. Liang, Y. Pan, Q. Guo, H. Hu, C. Wu, Q. Zhang, A novel analytical method for trace ammonium in freshwater and seawater using 4-methoxyphthalaldehyde as fluorescent reagent, J. Anal. Methods Chem. 2015 (2015) Article ID 387207. [78] M. Zhang, T. Zhang, Y. Liang, Y. Pan, Toward sensitive determination of ammonium in field: A novel fluorescent probe, 4, 5-dimethoxyphthalaldehyde along with a hand-held portable laser diode fluorometer, Sensor. Actuat. B: Chem. 276 (2018) pp. 356-361. [79] J. Růžička, E. Hansen, Flow injection analysis, Wiley, (1981) p. 207. [80] S.M. Gray, P.S. Ellis, M.R. Grace, I.D. McKelvie, Spectrophotometric determination of ammonia in estuarine waters by hybrid reagent-injection gas-diffusion flow analysis, Spectrosc. Lett. 39 (2006) pp. 737-753. [81] R.A. Segundo, R.B.R. Mesquita, M.T.S.O.B. Ferreira, C.F.C.P. Teixeira, A.A. Bordalo, A.O.S.S. Rangel, Development of a sequential injection gas diffusion system for the determination of ammonium in transitional and coastal waters, Anal. Methods 3 (2011) pp. 2049-2055. [82] S.M. Oliveira, T.I.M.S. Lopes, I.V. Tóth, A.O.S.S. Rangel, A multi-commuted flow injection system with a multi-channel propulsion unit placed before detection: Spectrophotometric determination of ammonium, Anal. Chim. Acta 600 (2007) pp. 29-34. 43

1043 1044 1045 1046 1047 1048 1049 1050 1051 1052 1053 1054 1055 1056 1057 1058 1059 1060 1061 1062 1063 1064 1065 1066 1067 1068 1069 1070 1071 1072 1073 1074 1075 1076 1077 1078 1079 1080 1081 1082 1083

[83] S.M. Oliveira, T.I.M.S. Lopes, I.V. Tóth, A.O.S.S. Rangel, Determination of ammonium in marine waters using a gas diffusion multicommuted flow injection system with in-line prevention of metal hydroxides precipitation, J. Environ. Monit. 11 (2009) pp. 228-234. [84] C. Henríquez, B. Horstkotte, V. Cerdà, A highly reproducible solenoid micropump system for the analysis of total inorganic carbon and ammonium using gas-diffusion with conductimetric detection, Talanta 118 (2014) pp. 186-194. [85] C. Henríquez, B. Horstkotte, V. Cerdà, Conductometric determination of ammonium by a multisyringe flow injection system applying gas diffusion, Int. J. Environ. Anal. Chem. 93 (2013) pp. 1236-1252. [86] J. Plant, K. Johnson, J. Needoba, L. Coletti, NH4-Digiscan: an in situ and laboratory ammonium analyzer for estuarine, coastal and shelf waters, Limnol. Oceanogr.: Methods 7 (2009) pp. 144-156. [87] L.O.C. Šraj, M.I.G.S. Almeida, I.D. McKelvie, S.D. Kolev, Determination of trace levels of ammonia in marine waters using a simple environmentally-friendly ammonia (SEA) analyser, Mar. Chem. 194 (2017) pp. 133-145. [88] P.-Y. Wang, J.-Y. Wu, H.-J. Chen, T.-Y. Lin, C.-H. Wu, Purge-and-trap ion chromatography for the determination of trace ammonium ion in high-salinity water samples, J. Chromatogr. A 1188 (2008) pp. 69-74. [89] T.Y. Lin, Y.T. Pan, H.Y. Lee, P.Y. Wang, C.H. Wu, Markedly enhanced purge-and-trap performance and efficiency for the determination of ammonium ion in high-salinity water samples, J. Chin. Chem. Soc. 59 (2012) pp. 718-726. [90] F.N. Ferreira, J.C. Afonso, F.V.M. Pontes, M.C. Carneiro, A.A. Neto, R.E. Junior, M.I.C. Monteiro, Ultrasound-assisted purge-and-trap extraction for simultaneous determination of low-molecular weight amines and ammonium in high salinity waters by ion chromatography, Microchem. J. 133 (2017) pp. 658-662. [91] Y. Zhu, D. Yuan, H. Lin, T. Zhou, Determination of ammonium in seawater by purge-and-trap and flow injection with fluorescence detection, Anal. Lett. 49 (2015) pp. 665-675. [92] W.S. Gardner, P.A. St. John, High-performance liquid chromatographic method to determine ammonium ion and primary amines in seawater, Anal. Chem. 63 (1991) pp. 537-540. [93] C.T. Kuo, P.Y. Wang, C.H. Wu, Fluorometric determination of ammonium ion by ion chromatography using postcolumn derivatization with o-phthaldialdehyde, J. Chromatogr. A 1085 (2005) pp. 91-97. [94] F.N. Ferreira, J.C. Afonso, F.V. Pontes, M.C. Carneiro, A.A. Neto, M.L. Tristao, M.I. Monteiro, Determination of low-molecular-weight amines and ammonium in saline waters by ion chromatography after their extraction by steam distillation, J. Sep. Sci. 39 (2016) pp. 1454-1460. [95] B. Pranaitytė, S. Jermak, E. Naujalis, A. Padarauskas, Capillary electrophoretic determination of ammonia using headspace single-drop microextraction, Microchem. J. 86 (2007) pp. 48-52. [96] S. Muniraj, C.-T. Yan, H.-K. Shih, V.K. Ponnusamy, J. Jen, Determination of ammonium in aqueous samples using new headspace dynamic in-syringe liquid-phase microextraction with in-situ derivitazation coupled with liquid chromatography-fluorescence detection, Anal. Chim. Acta 754 (2012) pp.54-60.

44

1084 1085 1086 1087 1088 1089

[97] I. Šrámková, B. Horstkotte, H. Sklenářová, P. Solich, S.D. Kolev, A novel approach to lab-in-syringe head-space single-drop microextraction and on-drop sensing of ammonia, Anal. Chim. Acta 934 (2016) pp. 132-144. [98] Y. Zhu, D. Yuan, J. Chen, K. Lin, Y. Huang, A novel ammonium-free seawater preparation method for determination of trace quantities of ammonium in seawater, Limnol. Oceanogr. Methods 16 (2018) pp. 51-56.

45

Table 1 Analytical features of spectrophotometric methods for ammonium determination in seawater from 1999-2019

Analytical method

Technique

Reagents

Chelating

λmax

Optical path

agents

(nm)

length (cm)

630

-

1-50 µM

640

3

Up to 428 nM

-

-

-

10-100 nM

EDTA

630

100

5 nM-10 µM

640

200

Up to 1000 nM

690

250

10 nM- 30 µM

Linear range

Precision

LOD

(% RSD)

(nM)

0.8 (10 µM)

Sample

Public

through

ation

-1

Ref.

put (h )

year

2100

-

2011

[54]

3.5

3

2011

[31]

-

-

2006

[30]

5.5

-

2015

[34]

5

30

2005

[32]

3.6

22

2014

[33]

360

24

2006

[55]

Manual IPB-phenol

operation-mi

Hypochlorite,

Trisodium

croplate

nitroprusside

citrate

DIC,

Trisodium

nitroprusside

citrate

reader IPB-phenol

SIA-SPE

IPB-phenol

GC-MS-SPE

IPB-phenol

IPB-phenol

IPB-phenol

IPB-phenol

DIC, nitroprusside

SFA-GD-L

Hypochlorite,

WCC

nitroprusside

SFA-LWCC

FIA-LWCC

LFA

DIC, nitroprusside DIC, nitroprusside DIC, nitroprusside

Trisodium citrate and EDTA

5.7 (44.6 nM) 3.8 (100 nM) 5 (10-100 nM)

Trisodium citrate and

4.4 (50 nM)

EDTA Trisodium citrate and EDTA

0.84-4.55 630

5

-

(0.18-1.1 µM)

IPB-salicylate IPB-1-naphtho l IPB-1-naphtho l

IPB-OPP IPB-OPP

rFIA FIA

Hypochlorite, nitroprusside DIC, acetone

Trisodium citrate

640

-

0.04-14.3 mM

735

1

Up to 286 µM

725

1

-

670

100/200

Up to 200 nM

700

3

Up to 100 µM

3

Up to 70 µM

700

3

Up to 200 µM

690

3

Up to 35 µM

0.7 (0.36

21000

32

2006

[56]

900

26

2009

[57]

200

-

2010

[25]

6/4

20

2015

[35]

200

3

2018

[36]

120

12

2018

[43]

150

12

2019

[44]

1.3

80

30

2018

[42]

mM) 0.6 (143 µM)

Manual operation-me

3 (0.3 µM)

DIC, acetone

EDTA

Hypochlorite,

Trisodium

nitroprusside

citrate

Manual

DIC,

Trisodium

operation

nitroprusside

citrate

DIC,

Trisodium

700/6

nitroprusside

citrate

00

DIC,

Trisodium

nitroprusside

citrate

DIC,

Trisodium

nitroprusside

citrate

OPA, sulfite

-

550

1

100-700 µM

-

7000

8

2016

[46]

None

543

1

Up to 57 µM

4.9

1900

10

2002

[52]

mbrane filter SFA-LWCC

IPB-OPP

iSEA

IPB-OPP

iSEA

IPB-OPP

rFIA

<4 (100 nM) 0.64-1.7 (10-50 µM) 0.23-3.36 (0-20 µM) 0.32-2.2 (2-20 µM)

Reaction of NH3-OPA-sulf

FIA

ite Hypobromite oxidation

Hypochlorite, FIA

potassium bromide

Table 2 Analytical features of fluorescence methods for ammonium determination in seawater from 1999-2019

Chemistry

OPA-sulfite

Technique

Buffer

Manual

Sodium

operation

tetraborate

Linear

Precision

LOD

(Ex/Em, nm)

range

(% RSD)

(nM)

-

<31

-

Seawater

35

-

-

Not mentioned

2

12

Ice core

Not mentioned

30

9

Saline

<2% (5-35);

water

-9% (<5)

7

30

Seawater

-

1000

120

Seawater

-

60

~90

360/420

Potassium OPA-sulfite

FIA

dihydrogen

365/-

phosphate Potassium OPA-sulfite

CFA

dihydrogen

360/420

phosphate OPA-sulfite

FIA

Sodium tetraborate

370/418-700

Potassium OPA-sulfite

FIA-GD

dihydrogen

310/390

phosphate OPA-sulfite

SIA

OPA-sulfite

SIA

Sodium tetraborate Sodium tetraborate

Sample

Wavelength

365/425 365/425

Up to 50 µM 0.2-60

1.7 (200

µM

nM)

Up to 5

3.8 (500

µM

nM)

Up to

~1 (0.5-4

100 µM

µM)

Up to 4

5.7 (800

µM

nM)

Up to 20 µM Up to 16 µM

through -1

put (h )

Type of

Salinity

Primary amines

sample

interference

interference

Coastal water

~3-5%

Publi cation

Ref.

year 1999

[62]

Not mentioned

1999

[64]

Not mentioned

2001

[65]

Negligible

2001

[66]

No effect

No effect

2005

[27]

No effect

Not mentioned

2006

[67]

Not mentioned

Not mentioned

2007

[68]

Manual OPA-sulfite

operation-m icroplate

Sodium tetraborate

360/430

OPA-sulfite with

CFA or FIA

None

365/425

formaldehyde OPA-sulfite

Modified

with

SIA (termed

formaldehyde

as ABA)

OPA-sulfite with formaldehyde

None

365/425

None

365/425

0.05-10

2.2 (>0.3

µM

µM)

0.1-12 µM

Matrix effect 5

0.6 (200

µM

nM)

Negligible

3600 1.1

nM)

0.005-25

Seawater

2007

[69]

(CFA);

2008

[7]

Correction is Seawater

No effect

8 (FIA)

needed at low levels

1

8

Seawater

Not mentioned

Not mentioned

2011

[71]

10

4

Seawater

Not mentioned

Not mentioned

2013

[72]

2011

[73]

Modified ABA with a portable

0.05-10 µM

0.3 (2 µM)

detector Sensitivity to

OPA-sulfite

should be corrected

2.2 (200 nM); 6.7 (1

16

CFA

Sodium tetraborate

370/427

0.05-25 µM

1-4 (5 nM-25 µM)

salinity <5

12

Seawater

variations especially at low levels

Interference at a low amine level was negligible; at a high level, the signal was depressed

Multi-pump ing flow OPA-sulfite

analysis with a rFIA

A slight Sodium tetraborate

365/425

Up to 16 µM

increase in <2 (5 µM)

13

32

Seawater

salinity of

concept OPA-sulfite with formaldehyde OPA-sulfite with formaldehyde OPA-sulfite with formaldehyde MOPA-sulfit e with formaldehyde

sensitivity with

<4%; 7.2% with uric acid

2011

[4]

~0.25% per 1%

Flow

Sodium

batch-SPE

tetraborate

360/425

1.67-300

3.5 (20

nM

nM)

Up to

0.8 (100

300 nM

nM)

0.032-15

3.2 (250

µM

nM)

0.25-1.2

2.35 (100

µM

nM)

Up to 5

3 (1000

µM

nM)

0.7

5

Seawater

Negligible

Negligible

2013

[74]

2.1

36

Seawater

Not mentioned

Not mentioned

2018

[75]

Not mentioned

Not mentioned

2014

[76]

Not mentioned

Not mentioned

2015

[77]

Not mentioned

Not mentioned

2018

[78]

FIA with a home-made

Sodium

portable

tetraborate

371.7/429.0

detector Manual

EDTA-Na

operation

OH

Manual

Sodium

operation

tetraborate

361/423

370/454

Natural 9.9

-

water, seawater Fresh

5.8

-

water, seawater

Manual operation MOPA-sulfit

with a

EDTA-Na

e

hand-held

OH

fluorescence detector

405/490

Fresh 3.5

-

water, seawater

Table 3 Analytical features of methods based on matrix separation for ammonium determination in seawater from 1999-2019 Separation technique GD

Technique

FIA-Fluorometry

Chemistry

OPA-sulfite pH indicator

GD

rFIA-Spectrophotometric

bromothymol blue

GD

GD

Multi-commuted FIA-Spectrophotometric Multi-commuted FIA-Spectrophotometric

pH indicator bromothymol blue pH indicator bromothymol blue

Linear

Precision

LOD

range

(%RSD)

(nM)

Up to 4 µM

5.7 (800 nM)

Up to 214

3 (7.1

µM

µM)

3.6-71.4 µM 3.6-71.4 µM

<1.5

Sample throughput (h-1)

7

30

643

135

3000

20

GD

system-Conductimetry

-

<2.0

1286

20

up to 2.0

µM);

bromothymol

µM

<2.5 (3

10

>8

Multi-syringe FIA-Conductimetry

-

2005

[27]

2006

[80]

2007

[82]

2009

[83]

2009

[86]

2011

[81]

2013

[85]

Estuarine water Surface and tap water

estuarine

and coastal water Transitional

5.5-55 µM

<2

1500

28

blue GD

Seawater

Ref.

Estuarine

pH indicator SIA-Spectrophotometric

year

water

µM) GD

Publication

sample

Seawater,

6 (1 Multi-pumping flow

Type of

and coastal water

4.2 µM-20 mM

<3

2500

32

Coastal water

GD GD

GD

Multi-pumping flow system-Conductimetry SFA-LWCC-Spectrophotometric Programmable flow-Spectrophotometric

0.5-25 µM

<1

5 nM-10

3.8 (100

µM

nM)

pH indicator

28

<1.2

bromothymol

nM-55.6

(<440

blue

µM

nM)

IPB-phenol

0.05-6.0

P&T

IC

-

P&T

IC

-

P&T

FIA-Fluorometry

OPA-sulfite

10-400 nM

IC

-

89-714 µM

Ultrasound-assisted P&T IC with a cation-exchange

Fluorometry

OPA-sulfite

column

µM 1.1 µM-5.6mM

0.05-5.0 µM

270

17

5.5

-

Coastal water Seawater

2014

[84]

2015

[34]

2017

[87]

Estuarine 15-440

20-40

and coastal water

<4

75/7.5

<4

Seawater

2008

[88]

<5

100/48

<2

Seawater

2012

[89]

7.4

4

Seawater

2015

[91]

~14000

<2

2017

[90]

2005

[93]

2007

[95]

2016

[94]

4.4 (200 nM) <10

Saline water Seawater,

<4

100

<4

photolyzed solution Human

Headspace-SDME

Capillary electrophoretic

-

5-100 µM

1500

3

blood, seawater and milk High

Steam distillation

IC

-

18-71 µM

≤4

2100

-

salinity water

Headspace dynamic in-syringe

LC with fluorescence detection

OPA-sulfite

LPME

0.625-10 µM

<6.6

330

<5

Lake water, river water

2012

[96]

2016

[97]

River Lab in syringe Headspace-SDME

Spectrophotometric

pH indicator

Up to 25

6 (10

µM

µM)

1800

17

water, coastal seawater

Highlights We explore the diverse range of methods available for the detection and analysis of ammonium in seawater to provide a basis for selection of the most suitable method. The developments of typically used methodologies in seawater analysis are summarized including ion-selective electrode, spectrophotometric, fluorometric and matrix separation methods. The main parameters assessed in the studies for ammonium analysis in seawater published in the last two decades (1999 to 2019) are also reviewed. Ammonium-free seawater preparation methods are summarized.