Preparation of Borax Cross-Linked Starch Nanoparticles for

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Preparation of borax crosslinked starch nanoparticles for improvement of mechanical properties of maize starch films Hao Lu, Na Ji, Man Li, Yanfei Wang, Liu Xiong, Liyang Zhou, Lizhong Qiu, Xiliang Bian, Chunrui Sun, and Qingjie Sun J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.8b06479 • Publication Date (Web): 21 Feb 2019 Downloaded from http://pubs.acs.org on February 22, 2019

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Journal of Agricultural and Food Chemistry

Preparation of borax crosslinked starch nanoparticles for improvement of

1

mechanical properties of maize starch films

2 3 4 5 6 7 8 9

Hao Lu1a, Na Ji1a, Man Lia, Yanfei Wanga, Liu Xionga, Liyang Zhoua, Lizhong Qiub, Xiliang Bianb, Chunrui Sunb, Qingjie Suna* a. College of Food Science and Engineering, Qingdao Agricultural University (Qingdao, Shandong Province, 266109, China) b. Zhucheng Xingmao Corn Developing Co., Ltd (Weifang, Shandong Province, 262200, China) 1

Equally-contributing authors

10

*Correspondence author (Tel: 86-532-88030448, e-mail: [email protected])

11

ABSTARCT: Recently, starch nanoparticles have attracted widespread attention

12

from various fields. In this study, a new strategy for preparing covalent-crosslinked

13

starch nanoparticles was developed using boron ester bonds formed between

14

debranched starch (DBS) and borax. The nanoparticles were characterized by Fourier

15

transform infrared spectroscopy (FTIR), transmission electron microscopy (TEM),

16

X-ray diffraction (XRD), dynamic light scattering (DLS), differential scanning

17

calorimeter

18

nanoparticles were spherical with a size of 100–200 nm. The formation of boron ester

19

bonds was confirmed by FTIR. The as-prepared starch nanoparticle exhibited a low

20

relative crystallinity of 13.6%–23.5%. Compared with pure starch film, the tensile

21

strength of starch film with 10% starch nanoparticles increased about 45%, and the

22

elongation at break percentage of starch film with 5% starch nanoparticles increased

(DSC),

and

thermogravimetric

analysis

ACS Paragon Plus Environment

(TGA).

The

obtained


Page 2 of 37

23

about 20%. The new strategy of forming starch nanoparticles by using boron ester

24

bonds will advance the research of carbohydrate nanoparticles.

25

KEYWORDS: debranched starch, short-chain amylose, boron ester bonds,

26

nanocomposite

27

INTRODUCTION

28

Starch is an abundant, inexpensive, and biodegradable polymer and is widely

29

applied in many areas. Recently, starch nanoparticles have aroused great interest in

30

academic research because of their nanoscale size, biodegradability, and

31

biocompatibility. Starch nanoparticles have been applied in a variety of areas, such as

32

in drug delivery and active material loading1,2, packaging materials3,4, and

33

emulsifiers5. Based on previous research, the methods for preparing starch

34

nanoparticles mainly include ultrasonication6,7, acid hydrolysis8, extrusion9,

35

nanoprecipitation10, enzymolysis11, and high-pressure homogenization12. Debranched

36

starch (DBS), mainly composed of linear glucan chains, is prepared by enzymatically

37

hydrolyzing the alpha-1,6-D-glucosidic bonds of amylopectin. DBS is an excellent

38

material for preparing starch nanoparticles. Recently, Sun et al. reported that starch

39

nanoparticles were obtained by combining enzymolysis with recrystallization

40

Size-controlled starch nanoparticles have been fabricated using DBS as raw material

41

13.

11.

42

However, there are few reports on the preparation of starch nanoparticles using

43

chemical crosslinking interactions. Reduction-sensitive starch nanoparticles were

44

prepared

via

the

reversed-phase

microemulsion


method

by

using

Page 3 of 37


14.

45

N,N-bisacryloylcystamine with the disulfide linkages

46

nanoparticles using POCl3 as a crosslinking agent

47

borax can form boron ester bonds with hydroxyl groups

48

reported that the adhesive layer of gelatinized starch paper was enhanced by borax

49

crosslinking

50

that it is feasible to prepare starch nanoparticles using borax as a crosslinker.

51

Moreover, we studied the influence of crosslinked starch nanoparticles on the tensile

52

properties, thermal stability, and other properties of maize starch films.

53

MATERIALS AND METHODS

54

Materials

17.

15.

Zhai et al. obtained starch

According to previous studies, 16.

Further, Shen et al.

Inspired by the interaction between starch and borax, we speculated

55

Waxy corn starch (98% amylopectin) and maize starch (26.5% amylose content)

56

were purchased from Tianjin Tingfung Starch Development Co., Ltd. (Tianjin,

57

China). Borax was purchased from Tianjin Guangcheng Chemical Reagent Co., Ltd.

58

(Tianjin, China). Pullulanase (E.C.3.2.1.41, 40 ASPU/g) was purchased from

59

Novozymes Investment Co. Ltd. (Bagsvaerd, Denmark).

60

Preparation of DBS

61

DBS was obtained using the strategy of Sun et al. (2014). Briefly, waxy corn

62

starch (5 g) was mixed with 50 mL of a phosphate buffer solution to obtain a

63

homogeneous suspension. Then, the starch dispersion was heated at 100 ° C for 30

64

min. After cooling the starch paste to 58 ° C, pullulanase was added, and the paste

65

was incubated at 58 ° C for 8 h. Subsequently, the pulluanase was inactivated by

66

heating the solution at 100 ° C for 15 min. Then, the precipitate was discarded by



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67

centrifuging (3500 rpm, 1 min). Four times the volume of ethanol was added into the

68

supernatant, and the pellets were obtained by centrifuging at 4000 rpm for 15 min.

69

After washing 3 times with water, the sediment was freeze-dried to obtain DBS.

70

Preparation of Starch Nanoparticles

71

Starch nanoparticles were obtained by borax crosslinking DBS. DBS (0.5 g or 1

72

g) was added into 50 mL of distilled water and heated in 120 °C for 5 min for

73

complete gelatinization. Then, 50 mL of borax solution (0.5 wt%, 1 wt%, 2 wt%) was

74

added dropwise to the DBS solution by continuously stirring using magnetic stirrer

75

(DF-101SZ, Kehua instrument equipment co. Ltd) at 600 rpm at 25 °C. At the end of

76

titration, the mixture was incubated for an extra 8 h at 25 ° C. Then, 100 mL of

77

ethanol was added, and the precipitate was collected by centrifuging at 3500 rpm for

78

15 min. The precipitate was washed several times and freeze-dried to obtain starch

79

nanoparticles. As a control, 50 mL of NaOH solution (pH=8.95) was added dropwise

80

to the DBS solution, and the following steps were the same as described above. The

81

obtained starch nanoparticles were denominated as DBSX-BY, where X and Y

82

indicates the concentration (w/v) of DBS and borax, respectively.

83

Preparation of Nanocomposite Films

84

The method of film preparation was referenced from Shi et al. (2013)

18,

with

85

some modifications. Maize starch (3.3 g) and 1.5 g of glycerol were mixed with 50

86

mL of distilled water. The starch dispersion was heated with vigorously stirring at

87

100 °C for 30 min. Then, the starch paste was cooled to 60 °C, and 25 mL of starch

88

nanoparticle suspensions (0 wt%, 5 wt%, and 10 wt% starch) was added, respectively,


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89

with stirring at 600 rpm. As comparison, 25 ml borax solution was added in the same

90

condition. The amount of borax added was consistent with the amount of borax in the

91

starch nanoparticles. After adding the nanoparticle suspensions or borax solution,

92

stirring of the mixture was continued at 600 rpm for 30 min. The film-forming

93

solution was degassed before casting. The mixture (20 mL) was evenly dispersed on

94

flat dishes (10 cm in diameter) and dried at 40 °C for 8 h. Finally, the starch films

95

were balanced at 75% relative humidity for 48 h at 25 °C.

96

Transmission Electron Microscope (TEM)

97

To observe the morphology of starch nanoparticles, the suspension, after 8 h of

98

reaction, was diluted 10 times with water for TEM measurement. The dilute solution

99

was ultrasonized at 25 KHz for 2 min. A drop of the diluted nanoparticles suspension

100

was added to a 300 meshes copper grid. The copper grid was freeze-dried at -40 °C.

101

The TEM analysis was measured using an HT7700 TEM (Hittach, Tokyo, Japan).

102

Dynamic Light Scattering (DLS)

103

The average size, size distribution, and polydispersity of the starch nanoparticles

104

were measured by DLS using a Malvern Zetasizer Nano (Malvern, Worcestershire,

105

UK). Samples were ultrasonically dispersed in ultrapure water (0.05%, w/v) and

106

analyzed at 25 °C 19.

107

X-ray Diffraction (XRD)

108

The crystal structure of the starch nanoparticles and starch nanocomposite films

109

were measured using an X-ray diffractometer (D8-ADVANCE, Bruker AXS Model,

110

Germany). The angle range was 5–40°.



111

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Fourier Transform Infrared Spectroscopy (FTIR)

112

The FTIR of the starch nanoparticles and starch films was measured using an

113

FTIR spectrophotometer (NEXUS-870; Thermo Nicolet Corporation, Madison, WI,

114

USA). The samples were scanned 64 times between 4000 and 500 with a spectral

115

resolution of 1 cm-1.

116

The Stability of Nanoparticles at Different pH Values

117

The starch nanoparticles (DBS2-B1, 0.05%, w/v) were ultrasonically dispersed in

118

ultrapure water, and the mean particle size and polydispersity index (PDI) were

119

analyzed at different pH values (pH=2, 4, 7, 8, and 10). For specific steps, refer to

120

section 2.6.

121

Thermal Properties of Nanoparticles

122

A differential scanning calorimeter (DSC; Mettler Toledo, Schwerzenbach,

123

Switzerland) was used to analyze the melting properties of the starch nanoparticles.

124

Approximately 3 mg of nanoparticles and 6 μL water were placed in an aluminum pan

125

and heated from 25 °C to 130 °C. The melting temperatures at the onset (To), peak

126

(Tp), and conclusion (Tc) were recorded. The enthalpy change of melting (ΔH) was

127

calculated based on the weight of dry samples.

128

The

thermal

stability

of

the

nanoparticles

was

analyzed

using

a

129

thermogravimetric analysis (TGA; Mettler Toledo, Greifensee, Switzerland). The

130

samples with a mass of about 3 mg were heated in pans from 30 °C to 600 °C.

131

The Water Resistance of Nanoparticles

132

The water resistance of starch nanoparticles was characterized by wettability and


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133

measured by an interface tensiometer (Sigma 700, Biolin). Native starch or starch

134

nanoparticles (1 g) was packed to a sample cell. Water was added in a beaker. Then,

135

the sample cell was suspended above the water surface. After the measurement was

136

completed, the contact angle (θ) was calculated.

137

Properties of Films

138

The thickness of the films was measured by a caliper. The opacity was measured

139

using an ultraviolet (UV)−visible Shimadzu 1601 PC spectrophotometer (Tokyo,

140

Japan) by scanning at 600 nm. The film bandings (1 × 4 cm2) were put into the quartz

141

cell. The opacity was obtained using the following equation:

142

Opacity = 𝑥

143

where A was the absorbance of the film at 600 nm, and x was the film thickness in

144

millimeters.

𝐴

(1)

145

The water vapor permeability (WVP) of the maize starch films was determined

146

using a weighing method. First, the films were balanced at 25 °C for 48 h in

147

desiccators with a relative humidity (RH) of 75%. Each film sample was sealed over a

148

conical flask (area = 12.56 cm2) filled with anhydrous calcium chloride (0% RH). The

149

quality of the conical flask was recorded every 12 h for 72 h. The WVP was

150

calculated using equation (2):

151

WVP =

152

where m is the weight increment of the films (g), d is the thickness (m) of each film,

153

A is the area of permeation (m2), t is the time lag for permeation (h), and P is the

154

pressure difference between the atmosphere containing calcium chloride and the

𝑚𝑑

(2)

𝐴𝑡𝑃



155

Page 8 of 37

distilled water-saturated environment.

156

The water solubility, moisture content, and swelling ratio of the films were

157

measured using a weighing method. The initial mass of the film bandings (1 × 4 cm2)

158

was recorded as m1. The dry mass (m2) of films dried at 105 °C for 24 h was

159

recorded. The mass of films immersed in distilled water at room temperature for 24 h

160

was named m3. Next, the film strips were dried at 105 °C for 24 h, after which the

161

mass of the dried sample was recorded (m4). The water solubility, swelling ratio, and

162

moisture content were calculated using equations (3), (4), and (5), respectively:

163

water solubility (%) =

164

swelling ratio (%) =

165

moisture content (%) =

𝑚2 ― 𝑚4

ⅹ100

𝑚4

𝑚3 ― 𝑚2 𝑚2

ⅹ100

𝑚1 ― 𝑚2 𝑚1

ⅹ100

(3) (4) (5)

166

The mechanical properties of the films were measured using a texture analyzer

167

(TAXT plus, Stable Micro Systems, Surrey, U.K.), fitted with an A/SPR probe.

168

Specific parameters were obtained from the literature 20. The tensile strength at break,

169

Young’s modulus, and elongation at break of the starch films were calculated using

170

the stress-strain curve.

171

TGA of the films was measured with a TGA (Mettler Toledo, Greifensee,

172

Switzerland). For specific steps, refer to section 2.10.

173

Scanning Electron Microscopy (SEM) of Films

174

Scanning electron micrographs were obtained using a scanning electron

175

microscope (S-4800, Hitachi Instruments Ltd., Tokyo, Japan). The films were placed

176

in liquid nitrogen and then fractured immediately. Films were sputter coated with gold


Page 9 of 37


177

at vacuum condition.

178

Statistical Analysis

179

Each experiment was repeated at least 3 times. The data were expressed using

180

mean values and standard deviations. The variance and significant differences were

181

analyzed by SPSS v.17.0 (SPSS Inc., Chicago, IL).

182

RESULTS AND DISCUSSION

183

Morphology of Starch Nanoparticles

184

The TEM images of the starch nanoparticles obtained from crosslinking DBS (1

185

wt%, 2 wt%) and borax (0.5 wt%, 1 wt%, and 2 wt%) are shown in Figure 1. The

186

shape of DBS1-B0.5 was irregular, and the size was 30–100 nm (Figure 1A). When

187

the concentration of borax was increased to 1% (DBS1-B1), the obtained particles

188

were aggregated with a size of about 50 nm (Figure 1B). As the concentration of

189

borax continued to increase (DBS1-B2), aggregation also increased (Figure 1C). The

190

results indicated that irregularly shaped nanoparticles could form at a 1%

191

concentration of DBS. As the DBS concentration increased to 2%, monodispersed

192

nanoparticles (DBS2-B0.5) were able to form with a size of 10–30 nm (Figure 1D).

193

Specially, DBS2-B1 (Figure 1E) showed a perfectly spherical structure, and the size

194

was in the range of 100–200 nm. However, a higher concentration of borax (2%) led

195

to aggregated nanoparticles. Combining the above experimental results, borax was

196

able to crosslink stretched DBS chains to form nanoparticles due to the boron ester

197

bonds formed between starch and borax. DBS contains a lot of hydroxyl groups.

198

Boron atoms in borax could form boron ester bonds with ortho-hydroxy groups in



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199

DBS to form starch nanoparticles. Similarly, starch–poly (vinyl alcohol) blends were

200

crosslinked by borax, enhancing their thermal stability. 21. In addition, Mohamed

201

reported that borax was a good crosslinking agent to improve the tensile and

202

modulus of starch film. We speculated that borax’s crosslinking ability was weak at

203

a low concentration (0.5%). However, excessive borax (2%) led to an aggregation of

204

nanoparticles. In addition, a DBS solution with a high concentration (2%) was

205

beneficial to the formation of nanoparticles.

22

206

We speculate that the morphology of nanoparticles was mainly affected by the

207

concentration of borax and DBS and the ratio of DBS to borax. When the

208

concentration of borax was 0.5%, the starch molecules were excessive. In this

209

condition, the crosslinking ability of borax was weak, so that the formed nanoparticle

210

structure was loose and the particles were dispersed. Figure 1 showed that the size of

211

DBS2-B0.5 was smaller than DBS1-B0.5. We hypothesized that borax molecule

212

crosslinked less starch molecules when the DBS concentration was 2%. Therefore, the

213

size of DBS2-B0.5 (Figure 1D) was smaller than DBS1-B0.5 (Figure 1A). In contrast,

214

when the concentration of borax was 1%, the crosslinking ability of borax was

215

enhanced. As the concentration of DBS was 1%, aggregation occurred between

216

nanoparticles due to excess borax crosslinks between particles (Figure 1B). However,

217

when the concentration of DBS was 2% and the concentration of borax was 1%, the

218

ratio of DBS and borax was proper. Borax molecules crosslinked an appropriate

219

number of DBS molecules and no obvious aggregation was observed between the

220

nanoparticles. In addition, spherical shape is more stable compared to other shapes,


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221

such as triangle and rectangle. Therefore, the morphology of DBS2-B1 was spherical

222

(Figure 1E). Obviously, when the concentration of borax was 2%, it had stronger

223

crosslinking ability compared with 0.5% and 1% borax. Therefore, the aggregations

224

of DBS1-B2 and DBS2-B2 were more obvious than other samples (Figure 1C and

225

Figure 1F).

226

Interestingly, although the ratio of DBS to borax in DBS1-B0.5 and DBS2-B1 was

227

the same, the morphology of the nanoparticles was different. Obviously, the

228

concentration of DBS2-B1 was higher than that of DBS1-B0.5. For DBS2-B1, there were

229

more DBS molecules crosslinked by boron ester bonds compared with DBS1-B0.5. We

230

speculate that sufficient cross-linked DBS molecules associated with each other to

231

form more perfect particles. Taken together, when the concentration of DBS and

232

borax was 2% and 1%, respectively, spherical nanoparticles with the size of 100–200

233

nm could be prepared.

234

Dynamic Light Scattering (DLS)

235

The hydraulic radius of the starch nanoparticles was determined using DLS

236

(Figure 2 and Table S1). As shown in Figure 2, the mean particle size range was

237

187.5 nm–266.9 nm, which was somewhat larger than that measured by TEM. During

238

DLS measurement, the hydraulic radius of the swelling starch nanoparticles was

239

measured. Moreover, nanoparticles may aggregate in aqueous environments, which

240

led to the increase in the size of the nanoparticles 23. Therefore, the size range of the

241

starch nanoparticles measured by DLS was larger than the range determined by TEM.

242

As observed in Table S1, when the concentration of borax was 2%, high PDI values



243

were observed for DBS1-B2 (0.55) and DBS2-B2 (0.51), indicating that particles were

244

aggregated at a high borax concentration. In contrast, the PDI of other nanoparticles

245

was about 0.18–0.25, which indicated that the nanoparticles were well dispersed. This

246

was consistent with the results of TEM.

247

FTIR of Starch Nanoparticles

248

The FTIR spectra (Figure 3A) of the starch nanoparticles revealed characteristic

249

vibrational bands corresponding to O–H stretching at 3300–3500 cm-1, and the peak

250

slightly shifted toward a longer wavelength. We speculated that the contents of the

251

hydroxyl groups were decreased due to the formation of covalent bonds with borax.

252

Nandkishore

253

consumed during the formation of covalent bonds with borax. Moreover, a

254

characteristic band of starch nanoparticles that appeared at 1339 cm−1 was attributed

255

to the stretching vibration of the B-O group. Similarly, Srinivas et al. reported that a

256

clear, obvious peak was detected at about 1338 cm-1, corresponding to the B–O

257

stretch

258

peaks at 1339 cm−1. The results indicated that the nanoparticles were formed by borax

259

crosslinking DBS.

260

The XRD of Starch Nanoparticles

24.

16

found that hydroxyl groups in the galactomannan backbone were

However, in the spectrum of blank control (DBS-NaOH), there were no

261

As shown in Figure 4, DBS exhibited a B-type crystal structure with

262

characteristic peaks at 16.9° and 22.9°. This was consistent with reports in the

263

previous literature 25. Moreover, a characteristic peak of the starch nanoparticles also

264

appeared at 16.9° with a low intensity, which indicated that the nanoparticles were


Page 12 of 37

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265

also B-type crystal. We speculated that the formation of boron ester bonds inhibited

266

the retrogradation of DBS due to the decreased contents of free hydroxyl groups in

267

DBS. Therefore, the relative crystallinity of the starch nanoparticles (13.6%–23.5%)

268

was lower than that of DBS (45.7%). Moreover, at the same concentration of DBS,

269

the higher the concentration of borax, the lower the relative crystallinity of the starch

270

nanoparticles.

271

The pH Stability of Starch Nanoparticles

272

The stability of the nanoparticles (DBS2-B1) at different pH values was measured

273

using DLS, and the results are shown in Figure 5. The nanoparticles exhibited a

274

bimodal distribution under acidic (pH = 2, 4) and neutral (pH = 7) conditions. The

275

first peak area was at about 7%. This indicated that the DBS2-B1 had undergone a

276

slight decomposition. We speculated that the boron ester bond was slightly destroyed

277

under acidic and neutral conditions. In contrast, DBS2-B1 had a single peak under an

278

alkaline condition (pH = 8, 10). Therefore, the size of DBS2-B1 was stable under an

279

alkaline condition. Similarly, Ding et al. reported that boron ester bonds were

280

dynamically reversible bonds that dissociated under acidic conditions and stabilized at

281

a pH around 9.5 26.

282

Thermal Properties of Starch Nanoparticles

283

As shown in Table 1, during the DSC measurement process, only DBS2-NaOH

284

and DBS2-B0.5 exhibited endothermic peaks with the ΔH of 12.13 J/g and 2.60 J/g,

285

respectively. Moreover, for other samples, there was no obvious endothermic

286

enthalpy in the DSC curves (data not shown). This indicated that borax was able to



287

inhibit the retrogradation of DBS. The results were in accordance with the XRD

288

results.

289

Figure 6 depicts the TGA and DTG of the starch nanoparticles. The thermal

290

degradation behaviors of the starch nanoparticles were mainly divided into two

291

processes. The mass loss of the first stage was due to water evaporation. The

292

degradation temperature of the nanoparticles and DBS2-NaOH was about 300 °C and

293

310 ° C, respectively (Figure 6A and 6B), and occurred in the second stage. This

294

indicated that the thermal stability of the starch nanoparticles was lower than that of

295

DBS. However, the weight loss of DBS1-B2 and DBS2-B2 was obviously lower than

296

DBS2-NaOH at 350–600 °C.

297

The Water Resistance of Nanoparticles

298

The contact angle (θ) of starch in water was 0 °. This indicated that starch

299

wettability was very high. However, the contact angle (θ) of starch nanoparticle was

300

17.36 °, indicating that the wettability of starch nanoparticles was much lower than

301

that of starch. This means that the starch nanoparticles have higher water resistance

302

than starch.

303

Physicochemical Analysis (WVP, Tensile Property, Swelling Property, Crystal

304

Structure, Surface Structure, Thermal Properties)

305

The thickness, WVP, and opacity of the maize starch films were measured (Table

306

S2). The thickness of the nanocomposite starch films was 115.67 μm–127.00 μm. The

307

opacity of the maize starch films was increased due to the addition of the starch

308

nanoparticles. The opacity of nanocomposite film with 10% starch nanoparticles was


Page 14 of 37

Page 15 of 37


309

about twice that of pure starch film. We guessed that the nanoparticles embedded in

310

the starch films prevented light transmission. Films with high opacity values could

311

protect products against light

312

values of the nanocomposite starch films were reduced with the raising concentration

313

of nanoparticles. Specially, the WVP value of starch films with 10% starch

314

nanoparticles decreased around 55% compared with pure starch films. The

315

enhancement of WVP was due to the increased compactness of the films contributed

316

by the starch nanoparticles 18.

27.

In addition, as can be seen in Table S2, the WVP

317

As shown in Table S3, there was no obviously difference in the moisture content

318

of the different starch films. The swelling ratio and water solubility, as important

319

properties of films, could provide insight into the behavior of films in an aqueous

320

environment. With the concentration of nanoparticles increased, the swelling ratio and

321

water solubility of films decreased. Compared to pure starch film, the swelling ratio

322

and water solubility of the films with 10% starch nanoparticles decreased around 11%

323

and 5.5%, respectively. We speculated that hydrogen bond interactions between the

324

starch matrix and starch nanoparticles enhanced the network of the film. Ortega et al.

325

also reported that the addition of silver nanoparticles decreased the water solubility of

326

nanocomposite films 27.

327

As shown in Table S4 and Figure 7, the addition of starch nanoparticles enhances

328

the strength and stretchability of the starch films. The tensile strength of

329

nanocomposite films with 10% starch nanoparticles (2.49 MPa) increased about 45%

330

compared to that of pure starch film (1.72 MPa). In addition, when 10% nanoparticles



Page 16 of 37

331

were added into the starch film, the Young's modulus increased about 72% compared

332

with that of pure starch film. The reason could be that starch nanoparticles in starch

333

films enhanced the starch network. Similarly, Ana et al. (2015)

334

films with acetylated starch nanoparticles enhanced the tensile strength of starch

335

films. Furthermore, the elongation at break of starch films was enhanced with the

336

addition of starch nanoparticles (Table S4). Compared to the pure starch film

337

(228.62%), the elongation at break of the nanocomposite film with 10% and 5%

338

starch nanoparticles increased 12% and 21%, respectively. When the amount of borax

339

added was 0.16% and 0.32%, the tensile strength of the borax crosslinked starch films

340

was higher than that of pure starch films. The tensile strength of borax crosslinked

341

starch films with 0.16% and 0.32% cross-linking agent (1.79 MPa and 2.08 MPa)

342

increased about 4% and 21% compared to that of pure starch film (1.72 MPa).

343

However, the tensile strength of nanocomposite films with 5% and 10% starch

344

nanoparticles was increased about 35% and 45%, respectively. This indicated that the

345

strengthening effect of starch nanoparticles was better than that of borax

346

cross-linking.

28

found that starch

347

Both the tensile strength and elongation at break of the nanocomposite film were

348

improved compared with that of pure starch film. We speculate that the high specific

349

surface area provided by nanoparticles caused stronger filler-matrix interfacial

350

interactions, which led to higher tensile strength of films reinforced with starch

351

nanoparticles. Moreover, the aldehyde groups of starch nanoparticles would destroy

352

inter-molecular hydrogen bonds between starch chains and enhance their mobility and


Page 17 of 37


353

then increased elongation at break value. Wu et al. also reported that agar would

354

destroy inter-molecular hydrogen bonds between polysaccharide chains and increased

355

elongation at break value29.

356

SEM Images of the Films

357

The SEM images of the nanocomposite films are shown in Figure 8. The surface

358

of the pure starch film had a smooth and homogeneous structure. In contrast, films

359

with 5% and 10% starch nanoparticles were observed as having rough and uneven

360

surfaces. Similarly, Dai et al. 30 reported that the surfaces of starch films incorporating

361

taro starch nanoparticles were rough. Furthermore, a small number of dispersed

362

nanoparticles were observed on the surface and in the cross-section of the starch films

363

with 5% starch nanoparticles. Moreover, there were obvious bulges and aggregated

364

nanoparticles on the surface of the nanocomposite film with 10% nanoparticles. It was

365

indicated that high concentrations of nanoparticles aggregated in the starch film.

366

López-Córdoba found that nanoparticles in cassava starch films showed a good

367

dispersion at a low concentration, while aggregated starch nanoparticles were found at

368

a high concentration. 31.

369

The FTIR and XRD of Films

370

The FTIR of starch films with different concentrations of starch nanoparticles are

371

presented in Figure S1. A strong vibration peak (O-H stretching) was observed at

372

around 3300 cm-1 in all films. In addition, the bending vibration of H2O appeared at

373

1644 cm-1. For pure starch film, a weak peak was detected at 2880 cm-1, which is

374

attributed to C-H stretching 32. However, the peak disappeared after the nanoparticles



375

were added. This indicated that the addition of nanoparticles weakened the vibration

376

caused by the C-H bond. In general, starch nanoparticles hardly changed the infrared

377

structure of the starch films.

378

As shown in Figure S2, for all the film samples, there were 2 peaks at around

379

17.0° and 20.0°, which indicated that the crystal structure of the films was the

380

B+V-type. Dai et al. also reported that corn starch films containing taro starch

381

nanoparticles were the B+V-type

382

formation of double-helixes 33. The peak 2θ at 20° corresponded to the single helical

383

crystal structure of the V-type, which indicated the formation of amylose–glycerol

384

complexes during processing

385

structure of the starch film was not affected by starch nanoparticles.

386

Thermal Properties of Films

34.

30.

The presence of peak 2θ at 17° indicated the

Moreover, the results indicated that the crystal

387

A TGA chart of the films was shown in Figure 9. The weight loss of the starch

388

films was roughly divided into 3 stages, which agreed with previous reports 35 (Figure

389

9A). As shown in Figure 9B, in the first step (35 -150°C), the evaporation of water led

390

to a decrease in the quality of the starch films. Then, the glycerol was degraded

391

between 150 and 250 ° C. In the final stage, the partially decomposed starch was

392

oxidized 36, 37. As shown in Figure 9B, the degradation temperature of nanocomposite

393

films with 5% (309.10 °C) and 10% (307.37 °C) starch nanoparticles was lower than

394

that of pure starch film (314.12 ° C). We speculate that starch nanoparticles with

395

lower thermal degradation temperatures (Figure 6) led to the easier degradation of

396

nanocomposite films compared with pure starch film.


Page 18 of 37

Page 19 of 37


397

In conclusion, we first prepared DBS nanoparticles using borax crosslinked DBS.

398

The morphology of DBS2-B1 was spheroidal, and the average diameter was about

399

100–200 nm. The presence of boron ester bonds (B-O) was detected using infrared

400

spectroscopy. The size and morphology of the nanoparticles were affected by the ratio

401

of DBS and borax. Nanoparticles with different structures could be prepared by

402

controlling the ratio of DBS to borax. The addition of starch nanoparticles enhanced

403

the tensile properties, toughness opacity, and WVP of maize starch films. This study

404

illustrated a new pathway for the fabrication of nanoparticles using boron ester bonds.



406

NOTES

407

The authors declare no competing financial interest.

408

ACKNOWLEDGMENTS

409

This work was supported by the National Key R&D Program of China (Project No.

410

2018YFD0400701), the National Natural Science Foundation, China (Grant No.

411

31671814), Major Agricultural Application Technology Innovation Project of

412

Shandong Province (Project No. SF1405303301), and Special Funds for Taishan

413

Scholars Project of Shandong Province (No. ts201712058).


Page 20 of 37

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414

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6. Liu, C. Z.; Li, M.; Ji, N.; Liu, J.; Xiong, L.; Sun, Q. J. Morphology and

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12. Shi, A.; Li, D.; Wang, L.; Li, B.; Adhikari, B. Preparation of starch-based

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13. Qiu, C.; Yang, J.; Ge, S. J.; Chang, R. R.; Xiong, L.; Sun, Q. J. Preparation and

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14. Yang, J.; Huang, Y.; Gao, C.; Liu, M.; Zhang, X. Fabrication and evaluation of

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Surface. B. 2014, 115, 368-376.

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15. Zhai, F.; Li, D.; Zhang, C.; Wang, X.; Li, R. Synthesis and characterization of

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polyoxometalates loaded starch nanocomplex and its antitumoral activity. Eur. J. Med


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17. Shen, J.; Zhou, X. F.; Wu, W. B.; Ma, Y, Q. Improving paper strength by gelation

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of native starch and borax in the presence of fibers. Bioresources. 2012, 7, 5542-5551.

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18. Shi, A. M.; Wang, L. J.; Li, D.; Adhikari, B. Characterization of starch films

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Polym. 2013, 96, 593-601.

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19. Liu, Q.; Li, F.; Lu, H.; Li, M.; Liu, J.; Zhang, S. L.; Sun, Q. J.; Xiong, L.

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Enhanced dispersion stability and heavy metal ion adsorption capability of oxidized

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starch nanoparticles. Food Chem. 2018, 242, 256-263.

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20. Yang, J.; Xiong, L.; Li, M.; Sun, Q. J. Chitosan-sodium phytate films with a

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one-step-consecutive-stripping and layer-by-layer-casting technologies. J. Agric.

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Food Chem. 2018, 66, 6104-6115.

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21. Sreedhar, B.; Sairam, M.; Chattopadhyay, D. K.; Rathnam, P. A. S.; Rao, D. V.

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M. Thermal, mechanical, and surface characterization of starch-poly (vinyl alcohol)

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blends and borax-crosslinked films. J. Appl. Poly Sci. 2005, 96, 1313-1322.

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22. Mohamed, R.; Mohd, N.; Nurazzi, N.; Siti Aisyah, M. I.; Mohd Fauzi, F. Swelling

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and tensile properties of starch glycerol system with various crosslinking agents. IOP

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23. Hebeish, A.; El-Rafie, M. H.; El-Sheikh, M. A.; El-Naggar, M. E. Ultra-fine

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characteristics of starch nanoparticles prepared using native starch with and without

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surfactant. J. Inorg. Organomet. P. 2013, 24, 515-524.

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24. Srinivas, G.; Burress, J. W.; Ford, J.; Yildirim, T. Porous graphene oxide

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frameworks: Synthesis and gas sorption properties. J. Mater Chem. 2011, 21,

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11323-11329.

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25. Kiatponglarp, W.; Tongta, S.; Rolland-Sabate, A.; Buleon, A. Crystallization and

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chain reorganization of debranched rice starches in relation to resistant starch

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formation. Carbohyd Polym. 2015, 122, 108-114.

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26. Ding, Q.; Xu, X.; Yue, Y.; Mei, C.; Huang, C.; Jiang, S.; Wu, Q.; Han, J.,

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Nanocellulose-mediated electroconductive self-healing hydrogels with high strength,

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plasticity, viscoelasticity, stretchability, and biocompatibility toward multifunctional

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applications. ACS. Appl. Mater Inter. 2018, 10, 27987-28002.

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27. Ortega, F.; Giannuzzi, L.; Arce, V. B.; García, M. A. Active composite starch

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films containing green synthetized silver nanoparticles. Food Hydrocolloids. 2017,

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70, 152-162.

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28. Teodoro, A. P.; Mali, S.; Romero, N.; de Carvalho, G. M., Cassava starch films

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containing acetylated starch nanoparticles as reinforcement: physical and mechanical

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characterization. Carbohyd Polym. 2015, 126, 9-16.

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29. Wu, Y.; Geng, F.; Chang, P.; Yu, J.; Ma, X. Effect of agar on the microstructure

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and performance of potato starch film. Carbohyd Polym. 2009, 76, 299-304.

501

30. Dai, L.; Qiu, C.; Xiong, L.; Sun, Q. J. Characterisation of corn starch-based films


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502

reinforced with taro starch nanoparticles. Food Chem. 2015, 174, 82-88.

503

31. López-Córdoba, A.; Medina-Jaramillo, C.; Piñeros-Hernandez, D.; Goyanes, S.

504

Cassava starch films containing rosemary nanoparticles produced by solvent

505

displacement method. Food Hydrocolloids. 2017, 71, 26-34.

506

32. Shen, G. C.; Ichikawa, M. Methane hydrogenation and confirmation of CHx

507

intermediate species on NaY encapsulated cobalt clusters and Co/SiO2 catalysts:

508

EXAFS, FTIR, UV characterization and catalytic performances. J. Chem. Soc.

509

Fadaday Trans. 1997, 93. 1185-1193.

510

33. Zhong, Y.; Li, Y. Effects of glycerol and storage relative humidity on the

511

properties of kudzu starch-based edible films. Starch – Stärke. 2014, 66, 524-532.

512

34. Shi, R.; Liu, Q.; Ding, T.; Han, Y.; Zhang, L.; Chen, D.; Tian, W., Ageing of soft

513

thermoplastic starch with high glycerol content. J. Appl. Poly Sci. 2007, 103, 574-586.

514

35. Jiang, W.; Qiao, X.; Sun, K. Mechanical and thermal properties of thermoplastic

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acetylated starch/poly (ethylene-co-vinyl alcohol) blends. Carbohyd Polym. 2006, 65,

516

139-143.

517

36. Wilhelma, H.; Sierakowski M.; Souzab G.; Wypychc F., Starch films reinforced

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with mineral clay. Carbohyd Polym. 2003, 52, 101-110.

519

37. García, N. L.; Famá, L.; Dufresne, A.; Aranguren, M.; Goyanes, S. A comparison

520

between the physico-chemical properties of tuber and cereal starches. Food. Res. Int.

521

2009, 42, 976-982.



522

FIGURE CAPTIONS

523

Figure 1. The transmission electron microscope images of starch nanoparticles: (A)

524

DBS1-B0.5, (B) DBS1-B1, (C) DBS1-B2, (D) DBS2-B0.5, (E) DBS2-B1, (F) DBS2-B2.

525

Figure 2. Particle size distributions of starch nanoparticles: (A) DBS1-B0.5, (B)

526

DBS1-B1, (C) DBS1-B2, (D) DBS2-B0.5, (E) DBS2-B1, (F) DBS2-B2.

527

Figure 3. FTIR of starch nanoparticles (B: partial enlarged image).

528

Figure 4. The XRD of starch nanoparticles and DBS.

529

Figure 5. The size distribution of nanoparticles (DBS2-B1) at different pH values.

530

Figure 6. Thermogravimetric analysis curves (A) and derivative thermogravimetry (B)

531

of starch nanoparticles.

532

Figure 7. The strain-stress curve of starch films.

533

Figure 8. The SEM images of surface (A: pure starch film, C: nanocomposite film

534

with 5% starch nanoparticles, and E: nanocomposite film with 10% starch

535

nanoparticles) and cross-section (B: pure starch film, D: nanocomposite film with 5%

536

starch nanoparticles, and F: nanocomposite film with 10% starch nanoparticles) of

537

starch films.

538


539

of starch films.


Page 26 of 37

Page 27 of 37

540


Table 1. Thermal properties of starch nanoparticles. Sample DBS-NaOH DBS2-B0.5

To (°C) 96.14±0.56 96.53±0.45

Tp (°C) 111.12±0.84 110.88±0.75

Tc (°C) 123.47±0.85 120.43±0.63


ΔH (J/g) 12.13±0.23 2.60±0.41


A

Page 28 of 37

B

500nm

C

500nm

D

200nm

E

500nm

F

500nm

500nm

541

Figure 1. The transmission electron microscope images of starch nanoparticles: (A)

542

DBS1-B0.5, (B) DBS1-B1, (C) DBS1-B2, (D) DBS2-B0.5, (E) DBS2-B1, (F) DBS2-B2.


Page 29 of 37


A

B

30

25

25

20

20

Intensity (%)

Intensity (%)

30

15

10

15

10

5

5

0

0 100

200

300

400

500

100

200

size (nm)

300

400

500

size (nm)

C

20

40

D

35

15

25

Intensity (%)

Intensity (%)

30

10

5

20 15 10 5

0 100

200

300

500

600

size (nm)

E

20

400

100

20

15

200

300

400

size (nm)

F

15

Intensity (%)

Intensity (%)

0

10

5

10

5

0 0

100

200

300

400

500

0 100

200

size (nm)

300

400

500

size (nm)

543

Figure 2. Particle size distributions of starch nanoparticles: (A) DBS1-B0.5, (B)

544

DBS1-B1, (C) DBS1-B2, (D) DBS2-B0.5, (E) DBS2-B1, (F) DBS2-B2.



Page 30 of 37

A

DBS2-B2

Transmittance (%)

DBS2-B1 DBS2-B0.5 DBS1-B2 DBS1-B1 DBS1-B0.5 DBS-NaOH

4000

3500

3000

2500

2000

1500

1000

500

-1

Wavenumber (cm )

B

Transmittance (%)

DBS2-B2 DBS2-B1 DBS2-B0.5 DBS1-B2 DBS1-B1 DBS1-B0.5 DBS-NaOH

1400

1380

1360

1340

1320

-1

Wavelength (cm )

545

Figure 3. FTIR of starch nanoparticles (B: partial enlarged image).


1300


Relative Intensity

Page 31 of 37

DBS DBS2-B2 DBS2-B1 DBS2-B0.5 DBS1-B2 DBS1-B1 DBS1-B0.5 5

10

15

20

25

30

2θ (°)

546

Figure 4. The XRD of starch nanoparticles and DBS.


35

40

45


Page 32 of 37

pH=2 pH=4 pH=7 pH=8 pH=10

20 18 16 14

Intensity(%)

12 10 8 6 4 2 0 10

100

1000

Size (nm)

547

Figure 5. The size distribution of nanoparticles (DBS2-B1) at different pH values.


Page 33 of 37


A

DBS2-NaOH

100

DBS1-B0.5 DBS1-B1 DBS1-B2

Weight (%)

80

DBS2-B0.5 DBS2-B1 DBS2-B2

60

40

20

100

200

300

400

500

600

Temperature (°C)

B 0.0000

DBS-NaOH DBS1-B0.5 DBS1-B1 DBS1-B2

-1

Derive. mass (wt% °C )

-0.0005

DBS2-B0.5

-0.0010

DBS2-B1 DBS2-B2

-0.0015

-0.0020

-0.0025 100

200

300

400

500

600

Temperature (°C)

548


549

of starch nanoparticles.



Page 34 of 37

Pure starch films Nanocomposite films with 5% starch nanoparticles Nanocomposite films with 10% starch nanoparticles

2.8 2.6 2.4 2.2 2.0

Stress (Mpa)

1.8 1.6 1.4 1.2 1.0 0.8 0.6 0.4 0.2 0.0 0

100

200

Strain(%)

550

Figure 7. The strain-stress curve of starch films.


300

Page 35 of 37


A

B

100μm

100μm

C

D d

100μm

E

100μm

F

100μm

100μm

551

Figure 8. The SEM images of surface (A: pure starch film, C: nanocomposite film

552

with 5% starch nanoparticles, and E: nanocomposite film with 10% starch

553

nanoparticles) and cross-section (B: pure starch film, D: nanocomposite film with 5%

554

starch nanoparticles, and F: nanocomposite film with 10% starch nanoparticles) of

555

starch films.

100μm


100μm


Page 36 of 37


A 100

Weight (%)

80

60

40

20

0 100

200

300

400

500

600

Temperature (°C)

0.0004

B


0.0002 0.0000

-1

Deriv. mass (wt% °C )

-0.0002 -0.0004 -0.0006 -0.0008 -0.0010 -0.0012 -0.0014 -0.0016 -0.0018 -0.0020 100

200

300

400

500

600

Temperature (°C)

556


557

of starch films.


Page 37 of 37


Graphic for table of contents

Borax

Debranched Starch

Crosslinked Starch Nanoparticles


Preparation of Borax Cross-Linked Starch Nanoparticles for

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