Phenylboronic Acid Functionalized Adsorbents for Selective and

Subscriber access provided by EKU Libraries

Food and Beverage Chemistry/Biochemistry

Phenylboronic Acid Functionalized Adsorbents for Selective and Reversible Adsorption of Lactulose from Syrup Mixtures Mingming Wang, Fayin Ye, He Wang, Habtamu Admassu, Yinghui Feng, Xiao Hua, and Ruijin Yang J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.8b02152 • Publication Date (Web): 15 Aug 2018 Downloaded from http://pubs.acs.org on August 16, 2018

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 37

Journal of Agricultural and Food Chemistry

Phenylboronic Acid Functionalized Adsorbents for Selective and Reversible Adsorption of Lactulose from Syrup Mixtures

Mingming Wang1, 2, Fayin Ye3, He Wang4, Habtamu Admassu1, 2, Yinghui Feng1,2, Xiao Hua1, 2*, Ruijin Yang1, 2*

1. State Key Laboratory of Food Science and Technology, Jiangnan University, 214122 Wuxi, China 2. School of Food Science and Technology, Jiangnan University, 214122 Wuxi, China 3. College of Food Science, Southwest University, 400715 Chongqing, China 4. Jiyang College, Zhejiang Agriculture and Forestry University, Zhuji, Zhejiang 311800, China

*Correspond to: Dr. Xiao Hua and Dr. Ruijin Yang Xiao Hua: [email protected] (X. Hua) Ruijin Yang: [email protected] (R. Yang)

1

ACS Paragon Plus Environment


1

ABSTRACT：：Boronate affinity materials have been widely used for enrichment of

2

cis-diol molecules. In this work, phenylboronic acid functionalized adsorbents were

3

prepared via a simple and efficient procedure by grafting phenylboronic acid groups

4

onto amino macroporous resins. Elemental analysis has confirmed the successful

5

functionalization of AR-1M and AR-2M with approximately 2.17% and 0.73% weight

6

percentage of boron. Comparatively, AR-1M possessed higher lactulose adsorption

7

capacity ( , 84.78±0.95 mg/g dry resin) under neutral condition (pH=7), while

8

the introduced glutaraldehyde spacer arms on AR-2M resulted in excellent adsorption

9

selectivity (α~23), high adsorption efficiency (π~22%), and fast adsorption/desorption

10

rate. The purity of lactulose ( ) through pH-driven adsorption (pH 7-8) and

11

desorption (pH 1.5) can be effectively improved depending on the ratio of lactulose to

12

lactose. When lactulose: lactose ≥1:1, ~95% was achieved. No significant drop

13

in (>90%) was observed after ten-consecutive repeats. Results demonstrated

14

that the newly developed method may achieve satisfactory performance in lactulose

15

purification.

16 17

KEYWORDS: Lactulose, phenylboronic acid functionalized adsorbent, adsorption

18

selectivity, pH-responsive, recyclability

19 20 21 22 23 24 25 2


Page 2 of 37

Page 3 of 37


26

INTRODUCTION

27

As a valuable lactose-originated non-digestive disaccharide, lactulose has been

28

commonly used in pharmaceuticals, nutraceuticals and food industries due to its

29

well-documented prebiotic properties.1-3 Currently, lactulose can be produced by

30

chemical-based isomerization of lactose (industrial scale) or through enzymatic-based

31

synthesis (lab-scale) using β-galactosidase, Caldicellulosiruptor saccharolyticus

32

cellobiose 2-epimerase (CsCE), etc.4-8 However, undesirable by-products of

33

monosaccharides and residual lactose remained in the isomerization syrup mixture

34

will most often lead to rising the costs associated with downstream lactulose

35

purification, since high-purity lactulose is mandatory for its medical uses and food

36

applications, especially for those who are lactose intolerance.1, 4, 9

37

Purification processing is a key operation for high-purity lactulose production.

38

Although monosaccharide can be efficiently removed from raw lactulose syrup by

39

using membrane separation, the fractionation of lactulose and lactose from

40

isomerization mixture is a challenging task due to the complexity of the syrup and the

41

highly structural similarity among them. Recently, several separation technologies,

42

such as methanolic crystallisation procedure10,

43

(SC-CO2) with alcohol-type cosolvent technique12, pressurized liquid extraction

44

(PLE)13 and room temperature ionic liquids (ILs)14, 15 approach have been applied to

45

obtain high-purity lactulose. However, those methods, efficient as they are, may not

46

appropriate for industrial process due to their sophisticated and tedious procedures,

47

considerable energy and organic solvent consumption. Very recently, a new innovative

48

strategy for lactulose and oligosaccharide purification has been developed through

49

selective fermentation of monosaccharides and lactose with Saccharomyces cerevisiae

50

and Kluyveromyces marxianus.9, 16 However, the final product was still a mixture of

11

, supercritical carbon dioxide

3



51

lactulose and oligosaccharides.

52

Boronate affinity based technique has been extensively used in the selective capture

53

and enrichment of cis-diol molecules (carbohydrates, nucleosides, glycoconjugates

54

and etc) from complex samples.17-23 The high selectivity inherits from the unique

55

chemical properties of boronic acids, which can reversibly form complexation with

56

cis-1,2- or cis-1,3-diols to generate 5- (stable) or 6-membered (less stable) cyclic

57

esters.17, 24 Furthermore, lactulose with typical cis-1,2-diol units shows higher binding

58

affinity than lactose with boronic acid groups (Supplementary Figure 1), which would

59

facilitate the separation of lactulose and lactose.25 To promote solid phase separation

60

and enhance the reusability of boronic acid groups, it generally requires immobilizing

61

this functional group on insoluble solid supports.26 Macroporous adsorption resins

62

(MARs) with high specific surface areas, large pore volumes (adsorption capacity),

63

regular and tunable pore sizes, and low cost, have been studied extensively and used

64

widely as excellent supports for industrial adsorption.26-28 Moreover, MARs generally

65

possess stable and interconnected frameworks with active pore surfaces, which make

66

them possible for further modification or functionalization with functional groups.26

67

The grafting of functional boronic acids onto MARs may somewhat favors the

68

selective capture of target lactulose from isomerization mixtures.

69

In the present work, a simple and effective modification procedure for the synthesis of

70

phenylboronic acid functionalized adsorbents (AR-1M and AR-2M) was established

71

by grafting phenylboronic acid groups onto amino macroporous resins (AR-0).

72

Physical characterization and adsorption/desorption properties (adsorption/desorption

73

kinetics and adsorption selectivity) of phenylboronic acid functionalized adsorbents

74

towards lactulose from isomerisates were investigated. The feasibility of adsorbent

75

recyclability and stability in ten consecutive capture/release cycles was also explored. 4


Page 4 of 37

Page 5 of 37


76

MATERIALS AND METHODS

77

Materials. Aminated poly (styrene-co-divinylbenzene) resin (AR-0, ~6.4 mmol/g dry

78

resin of -NH2 functional groups) was kindly donated by Tianjin Nankai Hecheng S&T

79

Co., Ltd. (Tianjin, China). 3-Aminophenylboronic acid (APBA, purity 98%),

80

p-formylphenylboronic acid (p-FPBA, purity 95%), sodium cyanoborohydride

81

(NaBH3CN, purity 95%), N,N-dimethylformamide (DMF, purity 99.8%) and HPLC

82

grade (purity>98%) of lactulose, lactose and epilactose were purchased from

83

Sigma-Aldrich (Shanghai, China). All other chemicals were of analytical grade and

84

obtained from Sinopharm Chemical Reagent Co. Ltd. (Shanghai, China).

85

Preparation of Phenylboronic Acid Functionalized Adsorbents. Two synthetic

86

protocols were used to functionalize poly(styrene-co-divinylbenzene) resins with

87

phenylboronic acid groups as depicted in Fig.1. Before functionalization, AR-0 was

88

first swelled thoroughly with DMF for 24 h. Then for route one, p-FPBA was

89

covalently bound to AR-0 via the formation of Schiff’s base between the formyl

90

groups of p-FPBA and the amino groups on the surface of AR-0.22, 23, 26 As to route

91

two, AR-0 was activated first by grafting of glutaraldehyde, and then the activated

92

AR-0 was chemically modified with APBA. Finally, sodium cyanoborohydride power

93

was added portionwise to reduce the Schiff’s base and the phenyboronic acid

94

functionalized adsorbents AR-1M (route 1) and AR-2M (route 2) were obtained after

95

washed with DMF, ethanol and deionized water and air-dried.

96

Characterization of Phenylboronic Acid Functionalized Adsorbents. Fourier

97

transform infrared spectroscopy (FT-IR) measurements of AR-0, AR-1M and AR-2M

98

were carried out on a Thermo Nicolet IS10 spectrometer (Nicolet, USA). X-ray

99

photoelectron spectroscopy (XPS) measurements for elemental analysis (C, N and B)

100

were performed using an AXIS Ultra DLD spectrometer (Shimadzu/Kratos, Japan). 5



101

The surface morphology of AR-0, AR-1M and AR-2M was observed with scanning

102

electron microscopy (SEM, 5.0 kV, SU1510; Hitachi Co., Japan).

103

Selective Adsorption of Lactulose and Lactose. The densities of phenylboronic acid

104

groups (µmol/g dry resin) on AR-1M and AR-2M was determined by calculating the

105

content of p-FPBA (λmax 256 nm) and APBA (λmax 293 nm) on a UV-3600

106

spectrometer (Techcomp, Shanghai, China) before and after the formation of boronic

107

ester27, 28, respectively. Unless otherwise stated, the adsorption studies were conducted

108

as follows: 1.0 g (dry mass) of the phenyboronic acid functionalized adsorbents was

109

added to 25 mL of lactulose and lactose binary solution (Lu:La=1:1, ×5 mg/mL) in a

110

100 mL Erlenmeyer flask. The initial pH values of the syrup ranged from 6.0 to 10.0

111

were adjusted by 0.1 M HCl/NaOH solution. The mixture was then shaken (150 r/min)

112

at 10-45oC for 12 h and samples were withdrawn at varying predetermined times

113

intervals for further analysis.

114

The equilibrium adsorption capacity in adsorption experiment, (mg/ g dry resin),

115

was calculated according to Eq. (1): = − ×

1

116

where and are the initial and equilibrium substrate concentrations (mg/mL),

117

respectively. refers the volume of syrup solution (mL), and is the dry mass (mg)

118

of the adsorbents.

119

The adsorption capacity at contact time (t) in adsorption experiment, (mg/ g dry

120

resin), was calculated as: = − ×

2

121

where represents the substrate concentrations (mg/mL) at contact time (min/h).

122

Adsorption selectivity (α) and adsorption efficiency (π, %) of adsorbents towards 6


Page 6 of 37

Page 7 of 37


123

lactulose from Lu-La binary solution were quantitatively defined as Eqs. (3) and (4): α=

⁄ 3 ⁄

π % = 100 ×

1000 × 4 × 342.30

124

where and (mg/g dry resin) denote respectively the equilibrium adsorption

125

capacities of lactulose and lactose. and are the equilibrium concentrations

126

(mg/mL) of lactulose and lactose, respectively. represents the content of

127

phenylboronic acid groups (µmol/g dry resin) on adsorbents.

128

pH-driven Desorption of Lactulose and Lactose from Adsorbents. After the

129

adsorption equilibrium was reached, pH-driven desorption of lactulose and lactose

130

from adsorbents was carried out as follows: the adsorbate-loaded AR-1M/AR-2M was

131

filtered and then desorbed with 25 mL pre-treated deionized water (pH 1-5, adjusted

132

by 1M HCl/NaOH) in 100 mL Erlenmeyer flask. Subsequently, the flask was

133

incubated at 25oC on a rotary shaker (150 r/min) for 2 h. Samples were withdrawn at

134

certain time intervals for further HPLC analysis.

135

The desorption capacity of lactulose and lactose ( , mg/g dry resin) and desorption

136

ratio (D, %) were defined to assess the desorption efficiency: = ×

5

% = ×

× 100% 6

−

137

where is the concentrations (mg/mL) of lactulose and lactose in the desorption

138

solution. is the volume (mL) of the desorption solution.

139

Selective Capture of Sugars by Phenylboronic Acid Functionalized Adsorbents

140

from Syrup Mixtures. AR-1M and AR-2M were tested for selective adsorption of

141

lactulose, lactose, fructose, glucose and epilactose from four syrup solutions: (1) 7



142

lactulose-lactose binary mixture (Lu:La=1:5, 1:2.5, 1:1, 2:1, 3:1, 4:1 and 5:1, ×5

143

mg/mL); (2) fructose-glucose binary mixture (Fru:Glu=1:1, ×5 mg/mL); (3)

144

fructose-glucose-lactulose-lactose mixture (Fru:Glu:Lu:La=1:1:1:1, ×5 mg/mL); and

145

(4) enzymatic reaction mixture (Lu:Epi:La=6.35:1.58:4.0, mg/mL) by using CsCE.6, 25

146

The pH values of these four different syrup solutions were adjusted to 8.0 by adding

147

0.1 M NaOH/HCl. Adsorption and desorption experiments were performed in an

148

identical manner as described above. The aliquots were sampled at different time

149

intervals and sugar concentrations were determined with HPLC analysis.

150

Initial purity ( " , %) and desorption purity (" , %) were defined as Eqs. (7) and (8)

151

in order to quantitatively evaluate the adsorption and desorption selectivity of

152

AR-1M/AR-2M towards different sugars from syrup solutions. " % =

" × × 100% 7

" % =

" × × 100% 8

153

" and " refer the initial and desorption concentration (mg/mL) of certain sugar,

154

respectively. and are, respectively, the total amount (mg) of sugars in the

155

initial and desorption solutions.

156

Recyclability of Phenylboronic Acid Functionalized Adsorbents towards

157

Lactulose. The reusability of AR-1M/AR-2M in sugar capture was investigated in

158

ten-consecutive adsorption/desorption cycles. Briefly, 1.0 g of AR-1M/AR-2M (dry

159

mass) was added to 25 mL of Lu-La binary solution (Lu:La=1:1, ×5 mg/mL, pH 7.0

160

for AR-1M and pH 8.0 for AR-2M) in a 100 mL Erlenmeyer flask. The mixture was

161

shaken (150 r/min) for 2 h at 25oC, and then the adsorbents were filtered. The

162

supernatant was collected for further HPLC analysis. Next, desorption process was

163

performed as described above (pH 1.5, 25oC, for 2 h). The separated adsorbent was 8


Page 8 of 37

Page 9 of 37


164

washed three times by deionized water. After air-dried overnight, the adsorbent was

165

used for sugar adsorption once again. The above adsorption/desorption cycles were

166

repeated for ten consecutively times.

167

HPLC Method for Sugar Analysis. Quantification of sugars concentration was

168

performed on a Hitachi L-2000 HPLC system (Hitachi Co., Japan) equipped with an

169

RI detector, using Asahipak NH2P-50 4E chromatographic column (4.6 mm×250 mm;

170

Showa Denko K.K, Japan).29 CsCE reaction mixture (lactulose, epilactose and lactose)

171

was analyzed using a Waters Alliance e2695 HPLC system (Waters Co., USA)

172

equipped with an RI detector (Waters 2414; Waters Co., USA) and Shodex VG-50 4E

173

chromatographic column (4.6 mm×250 mm; Shodex, Tokyo, Japan).27, 28

174

Statistical Analysis. All data expressed in this study were reported as mean±SD

175

(standard deviation). Each value represents the mean for three independent

176

experiments performed in duplicate with average standard deviations not exceeding

177

5%. SPSS statistical software 22.0 (SPSS Inc., Chicago, IL, USA) was used to

178

perform the statistical analysis.

179

RESULTS AND DISCUSSION

180

Characterization of Phenylboronic Acid Functionalized Adsorbents. FT-IR

181

spectra of AR-0, AR-1M, p-FPBA (Fig.2a) and AR-2M, APBA (Fig.2b) were used to

182

verify the successful grafting of phenylboronic groups. N-H stretching vibration at

183

3440 cm-1, 1501 cm-1, 903 cm-1 and C-N stretching vibration at 1108 cm-1 in both

184

AR-0, AR-1M and AR-2M indicated the presence of amino groups.24, 25 In Fig.2a and

185

b, it was found that both p-FPBA and APBA showed a characteristic stretching

186

vibration at 1342 cm-1, which is corresponded to B-O stretching of the phenylboronic

187

groups.18, 30, 31 When compared with AR-0, the adsorption band around 1342 cm-1 was

188

emerged for both AR-1M and AR-2M. The obtained results could indirectly prove that 9



189

the phenylboronic groups have been successfully introduced to AR-1M and AR-2M

190

through the amidation reaction.21-23

191

Chemical bonding of phenylboronic groups onto AR-1M and AR-2M surface was

192

further confirmed by XPS analysis. As shown in Fig.3, N 1s signal appeared at ~400

193

eV was observed for both AR-0 (Fig.3a), AR-1M (Fig.3c) and AR-2M (Fig.3e), but

194

only AR-1M (Fig.3d) and AR-2M (Fig.3f) exhibited the typical B 1s peak observed at

195

184-194 eV. Specifically, the peak emerged at 190.04 eV was corresponded to the

196

typical trigonal sp2-hybridized neutral B-species of the phenylboronic acid.19, 31 These

197

data indeed confirmed the success coupling of phenylboronic groups onto adsorbents

198

through boronic ester formation method.30, 31

199

The morphologies of the adsorbents were characterized by SEM. It was observed that

200

AR-0 showed a compact structure with a smooth surface (Fig.4a), whereas the pure

201

resin sample exhibited a rough surface, which occurs due to solvent swelling effect

202

(Fig.4b). The rougher surface also stated that the inner amino groups were much more

203

exposed, thus enabling the subsequent amidation reactions. In addition to AR-0,

204

AR-1M (Fig.4c) and AR-2M (Fig.4d) also had a rough surface accompanying by

205

pores with well distributed tiny holes, while all the skeletons retained their porous

206

structure and showed nearly uniform particle size (~300 µm). The changes on the

207

surface morphology would be beneficial for grafting more phenylboronic acid groups

208

which could obviously promote sugars capture.

209

Adsorption Properties of Phenylboronic Acid Functionalized Adsorbents. The

210

adsorption capacities of AR-1M (Fig.5a) and AR-2M (Fig.5b) on lactulose ( )

211

and lactose ( ) were gradually increased with the increasing of phenylboronic

212

acid groups due to the reversible covalent interaction between phenylboronic acid

213

groups and cis-diol compounds. More importantly, as depicted in Fig.S1, the binding 10


Page 10 of 37

Page 11 of 37


214

affinity of phenylboronic acid for lactulose (4-O-β-D-galactosyl-D-fructose) and

215

fructose was much higher than that of lactose (4-O-β-D-galactopyranosyl-D-glucose)

216

and glucose because of the typical cis-1,2-diol unit supplied by lactulose and

217

fructose,25 which can be easily linked with boronic acid group and to form a stable

218

5-membered boronic ester ring.17, 24 This is evidenced in Fig.5 that the increasing in

219

was much higher compared with that of . The discordance in adsorption

220

performance for lactulose and lactose could obviously increase the adsorption

221

selectivity (α) according to Eq.(3) and can be a principle of separation lactulose from

222

complex syrup. Furthermore, as depicted in Fig.1, the residual amino groups (-NH2,

223

~3,800 µmol/g dry resin) of AR-1M could cause nonspecific adsorption21, besides, the

224

residual amino groups might also participate in the adsorption of cis-dilos (lactulose

225

and lactose) through the B-N coordination.21, 32, 33 Those obviously exert influence on

226

and α. Whereas for AR-2M, the adsorption performance was well maintained

227

because of the presence of the interaction between amino groups and glutaraldehyde.

228

Moreover, the introduced flexible chains of glutaraldehyde (spacer arms) on AR-2M

229

were obviously favorable for boronate esterification with cis-diol compounds.

230

Therefore, it can be seen from Fig.5 that AR-2M not only has a superior adsorption

231

selectivity but also displays a higher adsorption efficiency (α=7.64, π~15.4% at

232

phenylboronic acid content of ~750 µmol/g dry resin) than AR-1M (α=2.27, π~11.2%

233

at phenylboronic acid content of ~2,600 µmol/g dry resin).

234

Accordingly, the weight percentages of the major elements in the phenylboronic acid

235

functionalized adsorbents were listed in Table 1. The mass concentrations of boron in

236

AR-1M (~2600 µmol of phenylboronic acid groups) and AR-2M (~750 µmol of

237

phenylboronic acid groups) were ~2.17% and ~0.73%, respectively. Comparatively,

238

the weight percentage of boron in AR-1M was almost 3 times higher than that of 11



239

AR-2M. Such a drastic decline in the content of boron could be reasonably attributed

240

to the successful grafting of bifunctional glutaraldehyde (spacer arms) on AR-2M

241

(Fig.1, route 2), which may exist in two types: resin-NH2-glutaraldehyde-NH2-resin

242

and resin-NH2-glutaraldehyde-phenylboronic acid group. These observations and

243

explanations provide insight that phenylboronic acid was presumably the sole

244

functional group of AR-2M, which might result in high lactulose selectivity, but failed

245

to increase the lactulose adsorption capacity.

246

Adsorption pH is a critical factor for boronate affinity. As stated in previous reports,

247

the binding of boronic acid groups (usually phenylboronic acid groups) to cis-1,2-diol

248

compounds occurred in two steps: phenylboronic acid group was first transformed to

249

tetrahedral anionic form [-B(OH)4-] (sp3) at alkaline conditions, and [-B(OH)4-] (sp3)

250

subsequently bonded with cis-1,2-diol units to form a stable 5-membered cyclic ester

251

(Fig.1).17, 30, 34 Given that most boronic acids were generally weak acids having a pKa

252

value of 8-9 (pKa of 8.7-8.9 for phenylboronic acid), it means that at pH≤ the pKa

253

value, most of the boronic acids still existed in the form of trigonal [-B(OH)2] (sp2),

254

which cannot react with cis-diol groups.21, 24 Based on this principle, phenylboronic

255

acid functionalized adsorbents required a basic pH (usually the pH should be> the pKa

256

value of the phenylboronic acid ligand) for binding.35 This is in good agreement with

257

the results obtained from AR-2M. As described above, phenylboronic acid was the

258

sole functional group on AR-2M, thus of AR-2M was definitely pH-dependent

259

and dramatically increased from ~5 to ~85 (mg/g dry resin) when the pH increased

260

from 6.0 to 8.0 and then kept almost unchanged (Fig.6b). Interesting, of

261

AR-2M was almost constant regardless the pH applied, this further confirmed that the

262

phenylboronic acid was the sole functional group of AR-2M. However, for AR-1M

263

(Fig.6a), was significantly decrease from approximately 60 mg/g dry resin at 12


Page 12 of 37

Page 13 of 37


264

pH 6.0 to about 30-40 mg/g dry resin at pH 7.0-10.0. The reason for this decline may

265

due to the influence of the pH affecting residual amino groups (-NH2), which is

266

stronger for lactose (through nonspecific adsorption) than for lactulose (through

267

reversible covalent binding with phenylboronic acid groups). Moreover, no obviously

268

change on was observed as the pH increased from 6.0 to 10.0, this might be

269

reasonably ascribed to the influence of residual amino groups, which could form an

270

alkaline microenvironment around the surface of AR-1M. Besides, as illustrated in

271

Fig.1, the residual amino groups of AR-1M may also react with the trigonal [-B(OH)2]

272

(sp2) and to form a B-N coordination complex (regard as Wulff-type boronic acids).32

273

B-N coordination complex with a low pKa value was obviously favors the adsorption

274

of cis-diols under neutral pH conditions.21, 32, 33 This interesting phenomenon is a great

275

advantage compared with traditional boronate affinity materials (such as AR-2M),

276

because it allows the selective adsorption of lactulose under neutral condition, which

277

is more favorable for industrial applications. Further studies are needed to understand

278

the positive adsorption of cis-diols through the B-N coordination.

279

Fig.6c and 6d provides the effect of adsorption temperature on sugars capture. It was

280

shown that of AR-1M was significantly (p96%, respectively. The same trend was also observed in AR-2M. Such a rapid 16


Page 16 of 37

Page 17 of 37


364

reduction in could be largely attributed to the inhibitory effect of the load of

365

more lactulose molecules at higher ratio of lactulose to lactose, since the binding

366

affinity of phenylboronic acid for lactulose (cis-1,2-diol unit) is much higher than that

367

of lactose.25 However, Fig.S2 shows that both and increased with the

368

concentration of lactulose and lactose at fixed Lu-La ratio of 1:1, which further

369

confirming the decisive effect of the ratio of lactulose to lactose on lactulose selective

370

capture. Moreover, it can be seen from Table 2A that AR-2M exhibited a lower

371

but a much higher at fixed Lu-La ratio compared with AR-1M, demonstrating

372

again that the introduced glutaraldehyde spacer arms on AR-2M can enhance the

373

adsorption selectivity towards lactulose but failed to improve the adsorption capacity

374

due to the reduction in phenylboronic acid groups.

375

Structure difference between lactulose (4-O-β-D-galactosyl-D-fructose) and lactose

376

(4-O-β-D-galactopyranosyl-D-glucose), namely the fructose moieties of lactulose

377

with typical cis-1,2-diol unit is responsible for their different binding affinity with

378

boronic groups.17, 22, 24 This explanation can be further supported by using fructose

379

and glucose as the model substrates. Table 2B clearly shows that a higher binding

380

affinity towards fructose was observed as expected at 1:1 ratio of fructose to glucose,

381

which ultimately resulted in a much higher &' for AR-1M (79.17±3.25%) and

382

AR-2M (90.45±5.47%).

383

Considering the fact that commercial lactulose syrup produced from chemical based

384

isomerization is a mixture of monosaccharides (i.e. fructose and glucose) and

385

disaccharides (e.g. lactulose and lactose).1,

386

mixtures of fructose-glucose-lactulose-lactose at a ratio of 1:1:1:1 was formulated to

387

investigate the applicability of AR-1M and AR-2M to selective adsorption of target

388

lactulose from the chemical isomerisates. Apparently, selective enrichment of

4

Herein，a model syrup consisting in

17



389

lactulose and fructose from isomerisates by using AR-1M and AR-2M are presented

390

in Table 2C. As a result, &' and were remarkably increased from 25% to

391

43.47±1.25%, 38.83±2.09% for AR-1M and 45.35±2.97%, 45.44±2.02% for AR-2M,

392

respectively. More importantly, small molecules such as monosaccharides can be

393

efficiently separated from lactulose syrup by using nanofiltration (NF) due to different

394

molecular weight cut-offs,37, 38 thus the affect of fructose on the selective adsorption

395

of AR-1M and AR-2M towards lactulose can be remarkably diminished through prior

396

NF process, implying that both AR-1M and AR-2M are still satisfactory in purifying

397

real isomerisates.

398

Table 2D claims the selective enrichment effect of AR-1M and AR-2M towards

399

lactulose from real CsCE-catalyzed reaction mixture, which contains lactulose,

400

epilactose and residual lactose at the ratio of 6.35:1.58:4.00 (×g/L). was

401

considerably increased from ~53% to 75.75±1.43% for AR-1M and 85.95±0.59% for

402

AR-2M, respectively. More importantly, both of the loaded lactose and epilactose

403

expect for lactulose could be released from adsorbents by using deionized water

404

(pH~5.8) because of their weak binding strength with boronic groups and/or the

405

nonspecific adsorption with amino groups (-NH2).21 Therefore, a higher * could

406

be obtained for both AR-1M (91.58±1.50%) and AR-2M (>98%) after first desorbing

407

with deionized water. The above results exhibited a good practicability of AR-1M and

408

AR-2M for real lactulose-enriched syrup.

409

Recyclability of Phenylboronic Acid Functionalized Adsorbents towards

410

Lactulose. From the viewpoint of practical applications, regeneration and

411

recyclability of phenylboronic acid adsorbents are other important features. Thus the

412

reutilization potential of AR-1M and AR-2M was evaluated in ten-consecutive

413

pH-driven adsorption/desorption cycles. As illustrated in Fig.8, no dramatic loss of 18


Page 18 of 37

Page 19 of 37


414

lactulose adsorption capacity was observed and the recovery was still up to ~90%

415

after 10-repeats. The slight reduction in adsorption capacity was probably attributed to

416

the cleavage of some unreduced Schiff’s base (-CH=N-) (Fig.1) under desorption

417

acidic condition (pH96

>96

50.80±0.76

53.18±1.77

80.68±3.10

97.33±2.07

102.33±2.86

116.21±2.23

1.36±0.23

0.61±0.41

ND

ND

ND

ND

92.75±3.70

95.67±1.92

>98

>98

>98

>98

(B) fructose-glucose mixture (Fru:Glu=1:1, ×5 mg/mL). Fru：Glu &' (%) AR-1M &' (mg/g) ,- (mg/g) ()./) (%) * AR-2M &' (mg/g) ,- (mg/g) ()./) (%) *

1:1 50 68.50±2.49 56.49±3.54 79.17±3.25 37.25±3.01 8.91±0.76 90.45±5.47

27



(C) fructose-glucose-lactulose-lactose mixtures (Fru:Glu:Lu:La=1:1:1:1, ×5 mg/mL). Fru：Glu：Lu：La

1:1:1:1 25 46.79±1.95 34.18±2.27 37.15±2.57 4.42±0.96 43.47±1.25 9.19± ±1.76 38.83± ±2.09 8.51± ±0.87 27.30±3.40 7.01±0.59 32.84±2.47 0.67±0.28 45.35±2.97 5.18± ±0.39 45.44± ±2.02 4.04± ±0.24

&' , ,- , , AR-1M

AR-2M

(%) &' (mg/g) ,- (mg/g) (mg/g) (mg/g) ()./) ） * （%） 12) ()* （%）） ()+) ） * （%） ()+3 ） * （%） &' (mg/g) ,- (mg/g) 45 (mg/g) 46 (mg/g)

()./) ） * （%） 12) ()* （%）） ()+) ） * （%） ()+3 ） * （%）

(D) CsCE reaction mixture (Lu:Epi:La=6.35:1.58:4.00, ×mg/mL). Lu: Epi: La 6.35:1.58:4.00 (g/L) 53.23 (%) 97.74±3.95 AR-1M (mg/g) 14.78±1.13 789 (mg/g) (mg/g) 40.20±2.07 75.75± ±1.43 ()+) （ %）） * * 91.58±1.50 ()+) （ %）） * 98.68±2.88 AR-2M (mg/g) 10.17±1.34 789 (mg/g) 15.14±3.25 46 (mg/g) +) 85.95± ±0.59 ()* （%）） * >98 ()+) （ %）） * *: desorption purity of lactulose from AR-1M and AR-2M after first washed with deionized water (pH 5.8~6.0).

28


Page 28 of 37

Page 29 of 37


Figures Fig. 1.



Fig.2.


Page 30 of 37

Page 31 of 37


Fig.3.



Fig. 4. (a)

(b)

(c)

(d)


Page 32 of 37

Page 33 of 37


Fig. 5.



Fig.6.


Page 34 of 37

Page 35 of 37


Fig.7.



Fig.8.


Page 36 of 37

Page 37 of 37


Graphic for table of contents


Phenylboronic Acid Functionalized Adsorbents for Selective and

Recommend Documents