Accepted Manuscript

A modified two-point titration method for the determination of volatile fatty acids
in
anaerobic systems
Zhe-Xuan Mu, Chuan-Shu He, Jian-Kai Jiang, Jie Zhang, Hou-Yun Yang, Yang Mu

PII: S0045-6535(18)30680-5
DOI: 10.1016/j.chemosphere.2018.04.038
Reference: CHEM 21186

To appear in: ECSN

Received Date: 23 January 2018

Revised Date: 24 March 2018
Accepted Date: 6 April 2018

Please cite this article as: Mu, Z.-X., He, C.-S., Jiang, J.-K., Zhang, J., Yang, H.-Y., Mu, Y., A
modified two-point titration method for the determination of volatile fatty acids in anaerobic
systems,
Chemosphere (2018), doi: 10.1016/j.chemosphere.2018.04.038.

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 ACCEPTED MANUSCRIPT
1 A modified two-point titration method for the determination of volatile fatty

2 acids in anaerobic systems

4 Zhe-Xuan Mu1#, Chuan-Shu He1#, Jian-Kai Jiang1*, Jie Zhang1, Hou-Yun Yang1,

5 Yang Mu1,2

1
7 CAS Key Laboratory of Urban Pollutant Conversion, Collaborative Innovation

8 Centre of Suzhou Nano Science and Technology, Department of Chemistry,

9 University of Science & Technology of China, Hefei, 230026, China

2
10 Anhui Province Key Laboratory of Polar Environment and Global Change,

11 University of Science & Technology of China, Hefei, 230026, China

12

13

14

15

#
16 These authors contributed equally to this work.

17

18

19

20

21 *Corresponding author:

22 Dr. Jian-Kai Jiang, E-mail: jkjiang@ustc.edu.cn

23

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24 Abstract

25 The volatile fatty acids (VFA) concentration plays important roles in the rapid

26 start-up and stable operation of anaerobic reactors. It’s essential to develop a simple

27 and accurate method to monitor the VFA concentration in the anaerobic systems. In

28 present work, a modified two-point titration method was developed to determine the

29 VFA concentration. The results show that VFA concentration in standard solutions

30 estimated by the titration method coincided well with that measured by gas

31 chromatograph, where all relative errors were lower than 5.5%. Compared with the

32 phosphate, ammonium and sulfide subsystems, the effect of bicarbonate on the

33 accuracy of the developed method was relatively significant. When the bicarbonate

34 concentration varied from 0 to 8 mmol/L, the relative errors increased from 1.2% to

35 30% for VFA concentration at 1 mmol/L, but were within 2.0% for that at 5 mmol/L.

36 In addition, the VFA composition affected the accuracy of the titration method to

37 some extent. This developed titration method was further proved to be effective with

38 practical effluents from a lab-scale anaerobic reactor under organic shock loadings

39 and an unstable full-scale anaerobic reactor.

40

41

42

43 Keywords: volatile fatty acids, two-point titration, anaerobic digestion, weak acid

44

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45 1. Introduction

46 Anaerobic digestion, a mature and versatile biotechnology, has been widely

47 applied to organic waste treatment and biogas production around the world (Mu et

48 al., 2006; Madsen et al., 2011; Purser et al., 2014). In the practical applications, poor

49 operational management and deficient process monitoring often result in start-up

50 failure or low operating performance (Lutzhoft et al., 2014). In anaerobic systems,

51 volatile fatty acids (VFA) are regarded as good indicators. An appropriate VFA

52 concentration keeps pH in the normal range and then maintains the microbial activity

53 of different microorganisms (Ahring et al., 1995; Björnsson et al., 2001). The

54 accumulation of VFA could induce the pH decline, leading to unstable operation or

55 even failure of anaerobic reactor (Florencio et al., 1995). Therefore, a simple and

56 accurate determination method is crucial to monitor the VFA concentration in the

57 anaerobic systems. In general, the main VFA determination methods are distillation,

58 colorimetry, gas chromatography (GC) and titration (Ai et al., 2011). Compared with

59 time-consuming distillation, inaccurate colorimetry and expensive GC, titration is a

60 preferred method to determinate the VFA concentration (Buchauer, 1998;

61 Moosbrugger et al., 1993a).

62 Titration is the process of determining the amount of a solution with a

63 concentration that is known and needed to react completely with a number of

64 specific examples to be analyzed (Ahring et al., 1995; Björnsson et al., 2001).

65 The process of titration, reagents are added dropwise into the analyte, usually using

66 a burette. Reagents are standard solutions whose concentrations are well known by

67 standardization. The addition of reagents is carried out continuously until it is

68 achieved equivalent between reagents and analytes, this condition is called the

69 equivalence point (Buchauer, 1998).

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70 alkalinity. As a result, the potential errors induced by other weak acids like

71 phosphate were inevitable between two titration endpoints. Subsequently, a novel

72 titration method taking into account of different weak acid subsystems was proposed

73 to improve the accuracy (Lahav et al, 2002; Moosbrugger et al., 1993b). However,

74 its better accuracy depended largely on the exact knowledge of the weak acid

75 concentrations, which made it less applicable. Recently, a more complicated method

76 combining pH and electrical conductivity was proposed to improve the accuracy

77 (Charnier et al. 2016).

78 This work aimed at developing an accurate VFA titration method with simple

79 manipulation and calculation. The two-point titration method was modified through

80 reselection of appropriate titration endpoints. Based on the charge balance and

81 ionization equilibrium in the solution, the VFA concentration was expressed as a

82 simplified equation, which was only associated with the titrant consumption at

83 different titration endpoints and dissociation constant. The modified two-point

84 titration method was firstly evaluated with the standard VFA solutions. The effects of

85 different weak acid subsystems on the titration accuracy were also investigated.

86 After that, the developed titration method was further verified with the effluents

87 from a lab-scale anaerobic reactor suffering organic shock loadings and an unstable

88 full-scale anaerobic reactor.

89

90 2. Development of modified two-point titration method

91 In anaerobic systems, typical components including carbonate ion, bicarbonate

92 ion, phosphate ion, hydrogen phosphate ion, dihydrogen phosphate ion, hydrogen

93 sulfide ion, sulfide ion and ammonium ion exist ubiquitously. According to the

94 Hendersone-Hasselbalch equation, the ionized concentrations of these interfering

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95 components varying with pH values were illustrated in Fig. 1 (Lutzhoft et al., 2014).

96 It could be observed that their ability to accept protons decreased at a lower pH value,

97 except for the dihydrogen phosphate. As acetic acid was the major VFA component

98 in anaerobic systems, its pKa value (i.e., 4.75 at 25 ) was commonly used as the

99 dissociation constant of VFA (Buchauer, 1998; Møller and Ward, 2011).

100 In general, the titration endpoints should be located in the range of pKa ± 0.5

101 and the difference between two titration endpoints should be greater than 0.25 to

102 avoid significant calculation errors (Moosbrugger et al., 1993b). In addition, the

103 influence of buffer systems like bicarbonate and phosphate should be weakened as

104 much as possible in the range of two titration endpoints. As shown in Fig. 1, only

105 about 1.3% bicarbonate and 0.3% dihydrogen phosphate could be protonated when

106 pH varied from 4.25 to 4.75. Hence, it could be assumed that the ionized

107 concentrations of interfering components are nearly zero in the titration except for

108 acetate, if 4.20 and 4.70 were chosen as the titration endpoints in the modified

109 two-point titration method.

110 Subsequently, sample solution with initial volume V was titrated by

111 hydrochloric acid with a constant concentration Ca until pH0 4.70, the equation of

112 charge balance could be obtained:

113 [H+] 0+n[Mn+] 0=[VFA-] 0+[OH-] 0+m[Xm-] 0 (1)

114 where [ ] indicates the molarity, Mn+ and Xm- denote all metal ions and all anions

115 excluding hydroxide, respectively.

116 After the addition of hydrochloric acid, the sample volume increased to V0, Eq.

117 (1) multiplied by V0 became:

118 [H+] 0V0+n[Mn+] 0V0=[VFA-] 0V0+[OH-] 0V0+m[Xm-] 0V0 (2)

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119 The hydrochloric acid continued to titrate until pH1 4.20. The sample volume

120 became V1 with added volume VX. The equation of charge balance could be

121 reformulated:

122 [H+] 1+n[Mn+] 1=[VFA-] 1+[OH-] 1+m[Xm-] 1 (3)

123 Similarly, Eq. (3) multiplied by V1 became:

124 [H+] 1V1+n[Mn+] 1V1=[VFA-] 1V1+[OH-] 1V1+m[Xm-] 1V1 (4)

125 In general, the common metal ions present in anaerobic system would not

126 precipitate with chloride ions. Therefore, the amount of Mn+ was almost constant

127 before and after the titration. Moreover, the protonated bicarbonate and dihydrogen

128 phosphate could be neglected when pH decreased from 4.7 to 4.2. The increased Xm-

129 was mainly attributed to the addition of chloride ions, namely CaVX. Eq. (4) subtracts

130 Eq. (2) to obtain:

131 [H+] 1V1-[H+] 0V0=[VFA-] 1V1+[OH-] 1V1+CaVX-[VFA-] 0V0-[OH-] 0V0 (5)

132 In the range of pH0 4.70 to pH1 4.20, the VFA ions and molecules existed

133 simultaneously in the solution. The concentrations of VFA ion could be expressed as

134 follows according to the ionization equilibrium:

[VFA ]0 V0 TVFA /(1 ( H ) 0 / K VFA ) TVFA / f ( H ) 0

135 (6)

[VFA ]1V1 TVFA /(1 ( H )1 / K VFA ) TVFA / f ( H )1

136 (7)

137 where TVFA is the total mole number of VFA, KVFA is the dissociation constant of

138 VFA, (H+)0 denotes the activity of H+.

139 Substituting Eq. (6) and Eq. (7) into Eq. (5), the initial VFA concentration

140 could be expressed as follows:

141 (8)

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142

143 3. Materials and methods

144

145 3.1 Chemical reagents

146 Sodium bicarbonate, sodium acetate, sodium phosphate, sodium sulfide,

147 propionic acid, butyric acid, ammonium chloride and hydrochloric acid of analytical

148 grade were bought from Sinopharm Chemical Reagent Company (Shanghai, China).

149 Their concentrations used in the experiments are summarized in the Supplementary

150 Material (Table S1).

151

152 3.2 Lab- and full-scale anaerobic reactors

153 A lab-scale upflow anaerobic sludge blanket (UASB) reactor with a working

154 volume of 2.9 L was used to carry out organic shock loading experiments. A water

155 jacket was used to maintain the reactor at mesophilic condition (37 oC). The

156 inoculum sludge was obtained from an expanded granular sludge bed reactor for

157 starch treatment (Binzhou, China). The UASB reactor was fed with glucose at 5 g

158 COD/L and other nutrients shown as follows (mg/L): NaHCO3 5000, NH4Cl 400,

159 Na2SO4 80, KCl 52, MgCl2· 6H2O 72, CaCl2· 2H2O 10, MnCl2· 4H2O 0.8,

160 CoCl2· 6H2O 1.2, H3BO3 0.2, CuCl2· 2H2O 1.1, Na2MoO4· 2H2O 0.1, ZnSO4· 7H2O

161 3.2, FeSO4· 7H2O 3.2, NiCl· 6H2O 0.5. The hydraulic retention time was kept at 28 h

162 under stable conditions, resulting in an organic loading of 4.29 kg COD/(m3d).

163 3-fold and 6-fold organic shock loading on the UASB reactor were carried out for

164 one day, respectively, by increasing influent COD concentration and keeping other

165 operating parameters unchanged.

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166 The full-scale anaerobic reactor of 600 m3 (Fuyang, China) was fed with

167 chicken manure (approximately 3.0%-6.0% w/w) and operated under continuous

168 mode at a flow rate of 1.67 m3/h. The operating temperature was maintained at 30 oC

169 in summer and 15 oC in winter. In the absence of buffer systems, the operation was

170 unstable during daily sampling in a week. The biogas production was less than 60

171 m3/d and the methane content ranged from 35% to 55%. Considering the high VFA

172 concentration in the effluents, samples were diluted eight times with distilled water

173 prior to experiments.

174

175 3.3 Analysis

176 All the effluent samples were filtered with 0.22 µm microfiltration membrane

177 prior to VFA determination. The VFA concentration directly measured by GC

178 served as standard value. The detailed procedures and parameters referred to a

179 previous study (Zhang et al., 2012). The detailed composition of VFA was also

180 measured by GC. The concentrations of ammonia nitrogen, phosphate and inorganic

181 carbon were obtained through Aquakem 200CD photometric analyzer (Thermo

182 Fisher Scientific, USA). The VFA titration was carried out using a TitraLab TIM865

183 automatic potentiometric titrator (HACH, America), which was composed of a

184 burette, a titration vessel equipped with magnetic stirring and a pH electrode. 30-40

185 mL sample solution was added into the titration vessel without pretreatment and

186 titrated with 50 mmol/L HCl at a titration speed of 0.7 mL/min and a stirring speed

187 of 300 rpm. The solution volume was recorded as V0 at pH0 4.7 and V1 at pH1 4.2,

188 respectively.

189

190 4. Results and discussion

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191

192 4.1 Method verification with standard solutions

193 Firstly, acetate solutions at various concentrations ranging from 1 to 30 mmol/L

194 were used to evaluate the accuracy of the modified two-point titration method. The

195 VFA concentrations estimated by titration agreed well with the corresponding

196 standard values measured by GC (Fig. S1). The relative errors were within 5.5% and

197 the relative standard deviation varied from 0.25% to 6.52%, suggesting a good

198 accuracy and reproducibility, respectively (Table S2). These results indicate that the

199 modified two-point titration method could estimate the VFA concentrations with a

200 satisfactory precision in a single component system.

201 When a multiple components system containing acetate and propionic/butyric

202 acid at different proportions were titrated, the relative errors became larger,

203 particularly in presence of propionic acid (Table 1). Although the pKa values of

204 acetic acid, propionic acid and butyric acid were close, the relative errors decreased

205 considerably when the pKa value of propionic acid (i.e., 4.87 at 25 ) or butyric acid

206 (i.e., 4.82 at 25 ) replaced that of acetic acid in the calculation (Table 1). Hence, it

207 is important to choose a suitable dissociation constant for better VFA determination.

208 Nevertheless, the errors induced by the discrepancy of dissociation constant were

209 still acceptable considering the unknown component in most practical situations.

210

211 4.2 Effects of various weak acid subsystems

212 Generally, the weak acid subsystems that coexist with VFA in anaerobic

213 systems can influence the accuracy of titration method. Therefore, the effects of

214 main weak acids including bicarbonate, phosphate, ammonium and sulfide on VFA

215 determination were investigated. The preliminary experiments were carried out to

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216 predict the potential errors induced by single weak acid. The results show that the

217 titrant consumption for 3 mmol/L bicarbonate, phosphate, ammonium and sulfide

218 could titrate 0.16, 0.08, 0.02 and 0.03 mmol/L VFA, respectively.

219 The effect of bicarbonate on VFA titration was noticeable (Fig. 2a). At low

220 VFA concentration of 1 mmol/L, the relative errors increased from 1.2% to 30%

221 with an increasing bicarbonate concentration from 0 to 8 mmol/L. When VFA

222 concentration reached 5 mmol/L, the relative errors were within 2.0%. In theory, the

223 influence of bicarbonate on VFA titration was weak in the range of pH0 4.70 to pH1

224 4.20 (Fig. 1). In fact, bicarbonate could combine with hydrogen ions and form

225 carbonic acid during the titration. When a part of carbonic acid decomposed into

226 carbon dioxide and escaped to the atmosphere, the pH in the system would increase,

227 resulting in considerable positive titration errors (Anderson and Yang, 1992). Slow

228 mixing speed could only mitigate rather than eliminate this phenomenon. In general,

229 the bicarbonate concentration is about 10 mmol/L in the anaerobic systems (Lahav

230 and Morgan, 2004). At low VFA concentration, although the relative errors were

231 remarkable, the small absolute errors would not induce a wrong assessment of

232 anaerobic systems. When the VFA accumulated in the anaerobic systems, the

233 modified titration method could estimate the VFA concentration accurately and

234 provide a warning signal accordingly.

235 In the presence of phosphate, the difference between VFA concentration

236 estimated by titration and that measured by GC was relatively small (Fig. 2b). At

237 VFA concentration of 1 mmol/L, the relative error increased from 3.0% to 11.0%

238 with an increasing phosphate concentration from 0 to 6 mmol/L. When the VFA

239 concentration was 5 mmol/L, the relative errors fluctuated within 2.5%. As

240 phosphoric acid could not decompose into gas escaping from solution, the positive

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241 errors were completely attributed to the titrant consumption induced by the

242 phosphate in the samples. In the anaerobic systems, the phosphate concentration was

243 relatively low and almost unchanged (Lahav et al., 2002). Thus, the effect of

244 phosphate on the modified two-point titration method was expected to be weak.

245 The VFA concentrations, in the presence of different ammonium concentrations,

246 obtained from GC and titration were almost coincident (Fig. 2c). The relative errors

247 were within 3.0% under experimental conditions. As the pKa value of ammonium

248 (i.e., 9.25 at 25 ) is much higher than that of VFA, the effect of ammonium on the

249 VFA determination was negligible at pH < 8 (Fig. 1).

250 The sulfide concentration also had a slight impact on the VFA determination

251 (Fig. 2d). The relative errors at either VFA concentrations (1 and 5 mmol/L) were

252 within 8.0%, suggesting the tolerance of titration method to the sulfide. In contrast, it

253 was reported that the loss of sulfide would induce significant errors in the algorithm

254 with regard to the 8-point titration method (Lahav et al., 2002). Also, the initial

255 sulfide concentration in the samples should be less than 25 mg/L. Similar to the

256 bicarbonate subsystem, sulfide in the completely protonated form could escape to the

257 atmosphere, leading to an increase in pH during the titration. Nevertheless, this kind

258 of error could be neglected at the experimental range owing to the high pKa value of

259 hydrogen sulfide (i.e., 7.04 at 25 ).

260

261 4.3 Method verification with anaerobic reactor effluents

262 The practical wastewater is much more complex than the standard solution. The

263 effluents from a lab-scale anaerobic reactor suffering different organic shock

264 loadings and from an unstable pilot-scale anaerobic reactor were tested to further

265 validate the modified two-point titration method. It is noteworthy that the

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266 concentrations of bicarbonate, phosphate, ammonium and sulfide in the samples

267 were unknown.

268 At the initial period, the stable VFA concentration obtained by the titration was

269 approximately 2.8 mmol/L, while the values measured by GC were almost zero (Fig.

270 3). This difference might be induced by the ubiquitous existence of weak acid

271 subsystems in the reactor effluents. Once the reactor suffered 3-fold organic shock

272 loading, the VFA concentration varied dramatically a half day later (Fig. 3). In

273 general, the VFA concentrations obtained by the titration were consistent with the

274 results measured by GC (Fig. S2). It implies that the titration method was accurate

275 enough when the VFA concentration was higher than 3 mmol/L. In the end, the

276 reactor resumed stable operation and the VFA concentration fell to its original level.

277 After suffering the 6-fold organic shock loading, the VFA concentration could

278 not recover to the initial level even half a month later. It could be deduced that the

279 acidification led to the failure of reactor operation (Lahav and Morgan, 2004). The

280 VFA concentrations obtained by titration were principally in agreement with the

281 results measured by GC (Fig. S3). It is worth noting that the VFA concentrations

282 determined by the titration were lower than the GC values after the shock loading

283 (Fig. 4a). The difference reached the maximum at the highest VFA concentration.

284 This was mainly due to the change in concentration of VFA components (Fig. 4b). At

285 the beginning, acetic acid was predominant component in the VFA. After the organic

286 shock loading, the concentration of propionic acid increased dramatically and kept

287 about twice the concentration of acetic acid.

288 With regard to the effluents from the unstable pilot-scale anaerobic reactor, the

289 results of GC and titration coincided well at such high VFA concentrations (Fig. 5a).

290 Moreover, the difference of determined VFA concentration stemming from two

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291 distinctive methods became smaller, which was contributed to the increased acetic

292 acid content in VFA (Fig. 5b). Meanwhile, the propionic acid and butyric acid

293 content keep stable relatively. These results further corroborate the importance of the

294 VFA composition for the modified two-point titration method.

295 Compared with other simple titration methods, the modified two-point titration

296 method minimized the influence of weak acid subsystems by narrowing the interval

297 between two titration endpoints. It did not involve the whole alkalinity that was

298 commonly applied previously (Kapp, 1984; Ripley et al., 1986; Anderson and Yang,

299 1992). Although its accuracy is still not comparable to the 5-point and 8-point

300 titration methods, it shows advantages in the case of unknown buffer subsystems.

301 The higher accuracy of these complicated methods depended largely on the known

302 sample characteristics such as the electrical conductivity, the total dissolved solids,

303 the concentrations of inorganic phosphate and sulfide (Vannecke et al., 2015). When

304 the sample characteristics were undetermined, a simple titration method was better

305 than the complicated titration (Lutzhoft et al., 2014). In addition, the principles of the

306 modified two-point titration method were based on the charge balance and ionization

307 equilibrium, which were easier to understand than the buffer capacity (Van De

308 Steene et al. 2002; Charnier et al., 2016). Considering its simplicity, reliability and

309 cost-effectiveness, this modified two-point titration method show a great application

310 potential in VFA monitoring.

311

312 5. Conclusions

313

314 The present work established a simple two-point titration method with reliable

315 accuracy for the determination of VFA in anaerobic systems. The VFA

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316 concentrations determined by the modified titration method was consistent with that

317 measured by GC with standard VFA solutions. Among various weak acid subsystems,

318 the effect of bicarbonate on the VFA determination was relatively significant. Further,

319 the modified two-point titration method was proved to be effective with effluents

320 from the lab-scale anaerobic reactor under organic shock loadings and the unstable

321 full-scale anaerobic reactor. In addition, the VFA composition affected the accuracy

322 of the titration method to some extent. This method shows a great potential to

323 monitor the VFA concentrations in practical applications.

324

325 Acknowledgements

326 This work was supported by the Natural Science Foundation of China

327 (51608502, 51538012 and 51478446), and the Fundamental Research Funds for the

328 Central Universities.

329

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394 Table 1. Effects of VFA compositions on the accuracy of modified two-point titration method

Acetate Propionic acid Butyric acid VFA by GC VFA by titration Relative error VFA by titration Relative error
(%) (%) (%) (mmol/L) (mmol/L)1 (%)1 (mmol/L)2 (%)2
100 0 0 5.10 ± 0.55 4.89 ± 0.13 -4.00 5.28 ± 0.14 5.40
95 5 0 5.24 ± 0.10 4.94 ± 0.06 -5.80 5.22 ± 0.10 -0.44
90 10 0 5.24 ± 0.37 4.73 ± 0.05 -9.60 5.21 ± 0.06 -0.47
85 15 0 5.23 ± 0.06 4.58 ± 0.13 -12.40 5.07 ± 0.13 -3.08
80 20 0 5.13 ± 0.15 4.54 ± 0.03 -11.40 5.02 ± 0.06 -2.00
60 40 0 5.43 ± 0.12 4.75 ± 0.08 -12.40 5.25 ± 0.12 -3.21
50 50 0 4.95 ± 0.53 4.47 ± 0.34 -9.60 4.95 ± 0.37 0.02
95 0 5 4.91 ± 0.12 4.81 ± 0.03 -2.20 5.07 ± 0.14 0.19
90 0 10 5.09 ± 0.35 4.78 ± 0.11 -6.20 5.04 ± 0.11 -1.00
85 0 15 5.21 ± 0.23 4.82 ± 0.08 -7.60 5.08 ± 0.10 -2.48
80 0 20 5.05 ± 0.08 4.65 ± 0.10 -8.00 4.90 ± 0.11 -2.89
60 0 40 4.97 ± 0.10 4.61 ± 0.29 -7.20 4.86 ± 0.32 -2.23
50 0 50 5.10 ± 0.14 4.74 ± 0.59 -7.20 4.99 ± 0.61 -2.22
395

396 1-Calculated with the pKa value of acetic acid

397 2-Calculated with the pKa value of propionic or butyric acid

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398 Figure captions

399

400 Figure 1. Ionized concentrations of general buffer subsystems at different pH values

401 (The notations of PO43-, CO32- and S2- are superposed).

402

403 Figure 2. Effects of (a) carbonate, (b) phosphate, (c) ammonia and (d) sulfide on

404 VFA titration (The detailed data were shown in Table S3).

405

406 Figure 3. Variation of VFA concentrations measured by GC and titration in the

407 effluents from a lab-scale anaerobic reactor under 3-fold organic shock

408 loading.

409

410 Figure 4. (a) Variation of VFA concentrations measured by GC and titration, and (b)

411 variation of VFA compositions in the effluents from a lab-scale anaerobic

412 reactor under 6-fold organic shock loading.

413

414 Figure 5. (a) Variation of VFA concentrations measured by GC and titration, and (b)

415 variation of VFA compositions in the effluents from an unstable full-scale

416 anaerobic reactor.

417

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IP
R
418

419
C
420 Figure 1

421

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T
P
I
R
C
S
U
N
422

423
A
424 Figure 2

425

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IP
R
426
427
C
428 Figure 3

429

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P
I
R
C
S
U
N
A
M
D
TE
430

431
P
432 Figure 4

433

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I
R
C
S
U
N
A
M
D
TE
434
435 P
436 Figure 5

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A modified two-point titration method was developed to monitor VFA

concentrations.

The titration method was effective for standard solutions and practical effluents.

Effect of bicarbonate subsystems on the titration accuracy was non-negligible.

The variation of VFA composition affected the titration accuracy to some extent.