Rheological and interfacial properties of basil seed gum modified with octenyl succinic anhydride

Journal Pre-proof Rheological and interfacial properties of basil seed gum modified with octenyl succinic anhydride Hadi Hashemi Gahruie, Mohammad Hadi Eskandari, Mohammadreza Khalesi, Paul Van der Meeren, Seyed Mohammad Hashem Hosseini PII:

S0268-005X(19)30157-2

DOI:

https://doi.org/10.1016/j.foodhyd.2019.105489

Reference:

FOOHYD 105489

To appear in:

Food Hydrocolloids

Received Date: 22 January 2019 Revised Date:

27 October 2019

Accepted Date: 4 November 2019

Please cite this article as: Gahruie, H.H., Eskandari, M.H., Khalesi, M., Van der Meeren, P., Hosseini, S.M.H., Rheological and interfacial properties of basil seed gum modified with octenyl succinic anhydride, Food Hydrocolloids (2019), doi: https://doi.org/10.1016/j.foodhyd.2019.105489. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2019 Published by Elsevier Ltd.

After 42 days

After 30 min Basil seed gum (BSG) OSA

C (unmodified BSG) S0 (OSA:BSG weight ratio of 0) S1 (OSA:BSG weight ratio of 0.01) S3 (OSA:BSG weight ratio of 0.03)

1

Rheological and interfacial properties of basil seed gum modified with

2

octenyl succinic anhydride

3 4

Hadi Hashemi Gahruie a, Mohammad Hadi Eskandari a, Mohammadreza Khalesi a, Paul

5

Van der Meeren b, Seyed Mohammad Hashem Hosseini a,*

6

a

7

Shiraz, Iran

8

b

9

University, Coupure Links 653, B-9000 Gent, Belgium

Department of Food Science and Technology, School of Agriculture, Shiraz University,

Particle and Interfacial Technology Group, Faculty of Bioscience Engineering, Ghent

10

* Corresponding author.

11

E-mail address: [email protected] (S.M.H. Hosseini)

12 13 14 15 16 17 18 19 20 21 22 23

1

24

Abstract

25

This study aimed to evaluate rheological and interfacial properties of basil seed gum (BSG)

26

esterified with octenyl succinic anhydride (OSA) at different OSA:BSG weight ratios (WRs)

27

of 0, 0.01, and 0.03. The amounts of added OSA were analyzed by high performance liquid

28

chromatography (HPLC) and ion mobility spectroscopy (IMS). The high correlation

29

coefficient (R2=0.998) between the results of HPLC (0%, 0.277%, and 1.01%) and IMS (0%,

30

0.31%, and 0.97%), obtained at different WRs, indicated that IMS can be considered as an

31

alternative analytical technique for HPLC to determine the extent of modification. The

32

entropy (image information content) of scanning electron micrographs of lyophilized BSG

33

was decreased after modification and attributed to relative disappearance of spherical

34

particles and formation of a structure with higher integrity. A decrease in interfacial tension,

35

an increase in contact angle and molecular weight, and more negative values of zeta-potential

36

were recorded after modification. All dispersions showed shear-thinning behavior with an

37

increase in apparent viscosity after modification. The first-order stress decay with a non-zero

38

equilibrium stress was better than other models for predicting the thixotropic properties. The

39

dilute solution properties were better fitted with slope-based models than intercept-based

40

models. A dominant elastic behavior was observed in BSG dispersions and corresponding

41

BSG-stabilized emulsions and improved after modification. The OSA-modified BSG

42

exhibited an improvement in the emulsifying and foaming capacities and colloidal stability

43

over time. Emulsions prepared with modified gums showed a smaller droplet size. OSA-

44

modified BSG might be a good candidate for improving the long-term stability of emulsions.

45 46

Keywords: Basil seed gum; Interfacial properties; Octenyl succinate anhydride; Ion mobility

47

spectroscopy; Rheological properties

48

2

49

1. Introduction

50

Basil (Ocimum basilicum L.) is found in different regions of Asia, Africa, and Central

51

and South America. Basil seeds and its gum are usually used in many desserts and traditional

52

beverages as a source of dietary fibers and a therapeutic agent (Gahruie, Eskandari, Van der

53

Meeren, & Hosseini, 2019). The advantages of basil seed gum (BSG) include hydrophilicity,

54

biocompatibility, low production cost, edibility, and appropriate film forming and

55

viscoelastic properties (Gahruie, Ziaee, Eskandari, & Hosseini, 2017). Based on the

56

molecular weight (MW) characteristics, BSG is fractionated into two main fractions

57

including PER-BSG and SUPER-BSG. The former constitutes about 69% of total BSG with

58

a MW of 5980 kDa. The MW of SUPER-BSG is around 1045 kDa. BSG is mainly composed

59

of D-galactose, D-glucose, D-mannose, D-xylose, L-arabinose and L-rhamnose in the

60

approximate proportion of 25:25:10:15:15:5, respectively. The average amount of uronic (D-

61

galacturonic and D-mannuronic) acid residues is 6.51%. SUPER-BSG (13.39%) has higher

62

uronic acid content than PER-BSG (3.84%), resulting in more anionic character (Naji-Tabasi

63

& Razavi, 2017a).

64

BSG can be used as a novel hydrocolloid emulsifier in food formulations (Naji-Tabasi

65

& Razavi, 2017a). Osano, Hosseini-Parvar, Matia-Merino, and Golding (2014) studied the

66

emulsifying properties of BSG under the effect of its concentration and purification (protein

67

removal from BSG). A two-stage high pressure homogenization at 35/8 MPa led to the

68

formation of stable emulsions with monomodal droplet distribution and average droplet size

69

(d32) below 1 µm at 0.3% BSG concentration. Protein removal reduced the adsorption

70

properties to the oil-water interface and developed larger oil droplets; however, protein-

71

depleted BSG still developed stable emulsions as compared to other hydrocolloids like sugar

72

beet pectin. Therefore, the authors concluded that the mechanism behind the emulsifying

73

ability of BSG may not be solely attributed to the surface-active protein moiety, but could

3

74

also be ascribed to the hydrophobic character of the polysaccharide itself (Osano et al., 2014).

75

Hosseini-Parvar, Osano, and Matia-Merino (2016) studied the effect of pH, ionic strength and

76

heat treatment on the emulsifying ability of BSG. A decrease in pH (< 4) and an increase in

77

ionic strength (> 40 mM) reduced the zeta-potential of BSG-stabilized oil droplets and thus,

78

an increase in droplet size was observed. Heating at extreme pH and ionic strength reduced

79

the emulsifying ability of BSG. The inclusion of hydrophobic segments of BSG molecules

80

into the oil droplets was confirmed by confocal laser scanning microscopy. The gel-like

81

behavior was reported as the main cause of the physical stability of emulsions against phase

82

separation at harsh pH and ionic strength conditions (Hosseini-Parvar et al., 2016).

83

Since the hydrophobic character of BSG molecules has a significant contribution to the

84

gum interfacial properties; hydrophobic modification might enhance the surface activity.

85

Simple chemical modification of hydrocolloids, particularly using octenyl succinic anhydride

86

(OSA) and dodecenyl succinic anhydride (DSA), has received increasing attention in recent

87

years. Pan, Yang, and Qiu (2015), Sarkar, Gupta, Variyar, Sharma, and Singhal (2013) and

88

Sarkar and Singhal (2011) optimized the synthesis of OSA-modified gum Arabic and

89

reported that the functional performance was improved. An improvement in the emulsifying

90

properties of gum Arabic after esterification with DSA was also reported by Wang, Williams,

91

and Senan (2014). Recently, we reported the effect of BSG modification with OSA on the

92

characteristics of BSG-based edible films (Gahruie et al., 2019). However, to our knowledge,

93

the current work is the first report in which the physicochemical, rheological, and interfacial

94

properties of OSA-modified BSG have been evaluated under the effect of OSA:BSG weight

95

ratio.

96

2. Materials and methods

97

2.1. Materials

4

98

Basil seed gum (BSG) was obtained from Reyhan Gum Parsian (Tehran, Iran). As

99

stated by the supplier, the extraction process of BSG included: 1) removing foreign bodies in

100

excess amounts of ethanol; 2) drying at 45 ºC; 3) soaking in distilled water at 50 ºC; 4)

101

shearing and scrapping the hydrocolloids off the surface of swollen seeds; 5) filtration

102

through a fine cloth; and 6) drying. 2-Octen-1-ylsuccinic anhydride (OSA, 97% purity) and

103

egg albumin were purchased from Sigma-Aldrich (St. Louis, MO, USA). HPLC-grade

104

acetonitrile and methanol were purchased from Merck Co. (Darmstadt, Germany). Pure

105

canola oil was kindly donated by Narges Shiraz Oil Company (Shiraz, Iran). All other

106

chemicals were of analytical grade and used as received.

107

2.2. Modification of basil seed gum

108

BSG was modified with OSA using the method described by Pan et al. (2015) with

109

some modifications. BSG dispersion (1.5% w/v) was prepared in deionized water and

110

hydrated overnight. After that, the pH was adjusted to 8.0 using 0.5 M NaOH. Ethanolic

111

solution of OSA was added to BSG dispersion at 25 °C to reach different OSA:BSG weight

112

ratios (WR) of 0, 0.01 and 0.03. OSA and BSG were then allowed to react at 40 ± 2 °C for 90

113

min. The pH was maintained at 8.0. The reaction was terminated by the addition of 0.1 M

114

HCl to pH 6.0. Washing with absolute ethanol was performed three times to remove the

115

residual amounts of OSA. The precipitates were then freeze-dried. The modified samples

116

were named as S0, S1 and S3 indicating the WRs of 0, 0.01 and 0.03 during the modification,

117

respectively. There was a possibility that the washing step with ethanol (to remove free OSA

118

molecules) might had a purifying effect on the modified samples and hence led to different

119

amounts of hydrocolloids molecules at a same concentration of different samples. Table 1

120

reports the moisture, protein, and carbohydrate contents (%) of unmodified BSG (denoted as

121

C) and modified BSG (denoted as S0, S1 and S3) samples. The observed changes were not

122

significant.

5

123

2.3. Determination of the extent of modification

124

2.3.1. HPLC analysis

125

The amount of bound OS was determined according to the method described by Shi et

126

al. (2017) with slight modifications. Exactly, 0.1000 g of dry samples were immersed in 2

127

mL of NaOH (4 M) and stirred overnight. The alkali treated solutions (0.4 mL) were mixed

128

with 3.6 mL of 1 M HCl and then made to volume with acetonitrile in a 5 mL volumetric

129

flask. The samples were then analyzed using an HPLC system (Knauer, Germany) equipped

130

with Smartline pump 1000 and C18 column (sphere image 80-5 ODS 2, particle size 5 µm,

131

length 300 mm, internal diameter 4 mm, Knauer, Germany). A mixture of acetonitrile and

132

water (50:50 v/v) containing 0.1% formic acid was used as the mobile phase at the flow rate

133

of 0.8 mL/min. Samples were filtered through 0.45 µm syringe filters and then injected (10

134

µL). The UV absorption was measured at 205 nm using 2100 UV detector (Knauer,

135

Germany). The OS content was calculated from the OSA standard curve (y = 2739.4x +

136

113801) constructed by plotting the peak area (y) vs. OSA concentration (x, µg/mL). The

137

amount of bound OS (%) was calculated using Eq. 1.

138

%OS = (250Wt/W) × 100

139

where, W is the dry weight (g) of the sample, Wt is the OS content determined from the

140

standard curve, and 250 is the dilution factor.

141

2.3.2. Ion mobility spectrometry

(1)

142

A series of OSA standard solutions was prepared in ethanol and then analyzed using an

143

ion mobility spectrometer (IMS, model 300; TOF Tech. Pars Co., Iran). The standard curve

144

(y = 0.0069x + 0.0087) was then prepared by plotting the peak area (y) vs. OSA concentration

145

(x, µg/mL). Subsequently, the BSG was dissolved in ethanol at a concentration of 20% and

146

injected (5 µL). Detection and quantification of OS groups in modified gums using IMS was

147

carried out in positive mode. The optimized experimental conditions for obtaining the ion 6

148

mobility spectra were as follows: corona voltage 2500 V; drift voltage 8000 V; drift gas (air)

149

flow 380 mL min−1; carrier gas (air) flow 120 mL min−1; shutter grid 250 µs; cell temperature

150

200 °C; and injector temperature 260 °C.

151

2.4. Molecular weight

152

The molecular weight of modified BSG was determined using a dynamic light

153

scattering instrument (DLS, SZ100, Horiba, Japan) operating in the static mode (Scattering

154

angle 90°) at 25 °C. The stock dispersions (5, 2.5, 1.25, and 0.625 mg/mL) of samples were

155

prepared in deionized water. After complete hydration, centrifugation was performed at 1000

156

g for 3 min to remove possible impurities.

157

2.5. Zeta potential

158

The zeta potential of BSG dispersions (0.1% w/w) was determined using DLS (SZ100,

159

Horiba, Japan) at 25 °C. The electrophoretic mobility was converted to zeta potential using

160

the Smoluchowski equation.

161

2.6. Contact angle

162

BSG samples were shaped into small pellets with a smooth surface by a lab scale press.

163

The static sessile drop method was utilized to measure the contact angle (θ) of BSG using a

164

drop shape analyzer (DSA 100, KRÜSS GmbH, Hamburg, Germany). Pellets were placed at

165

the bottom of pure canola oil. A water droplet (2 µL) formed at the tip of the needle was

166

placed on the BSG pellets. A CCD camera equipped with macro lens was used to take

167

pictures of the drop shape. Analysis was performed by the DSA software.

168

2.7. Rheological properties of gum

169

2.7.1. Temperature effect on viscosity

170

The effect of temperature on the viscosity of modified gums was studied using a Rapid

171

Visco Analyser (RVA Starch Master 2, Perten, Australia). Briefly, 0.3 g of BSG was gently

7

172

mixed with 25 mL of distilled water in the RVA cup at room temperature. Changes in the

173

viscosity as a function of temperature were monitored using the following heating/cooling

174

profile: equilibration for 1 min at 50 °C, heating from 50 to 95 °C at 12.3 °C/min, holding for

175

2.6 min at 95 °C, cooling from 95 to 50 °C at 12.3 °C/min and holding for 2 min at 50 °C.

176

The rotation speed of the paddle was 160 rpm.

177

2.7.2. Apparent viscosity

178

The rheological behavior of unmodified and modified gums (0.3% w/v) was studied at

179

20 °C and the shear rate range of 1 to 62.4 s-1 using a rheometer (MCR 302, Anton Paar,

180

Graz, Austria) equipped with cone-plate geometry (type CP25-1, cone diameter = 25 mm,

181

gap size = 0.052 mm, and cone angle = 1˚). The temperature of the bottom plate was

182

controlled with a Peltier system (Viscotherm VT2, Phar Physica) with an accuracy of ±0.1

183

°C. The apparent viscosity was reported at the shear rate of 51.1 s−1. Shear stress vs. shear

184

rate data was fitted using four common models namely Power Law, Herschel-Bulkley,

185

Bingham, and Casson (Eqs. 2-5, respectively).

186

τ = kγn

(2)

187

τ = τ0 + kγn

(3)

188

τ = τ0 + µ γ

(4)

189

τ0.5 = τ00.5 + kγ0.5

(5)

190

where, γ is the shear rate (s-1); τ is the shear stress (Pa); τ0 is the yield stress (Pa); k is the

191

consistency coefficient (in Pa.sn for Herschel-Bulkley and Power Law and Pa.s0.5 for Casson);

192

n is the flow behavior index (dimensionless); and µ is the Bingham viscosity (Pa.s).

193

2.7.3. Time dependency

194

BSG dispersion (0.3% w/v) was subjected to the constant shear rate (γ) of 200 s-1. Shear

195

stress (τ) was then recorded as a function of shearing time (t) until reached to the steady state.

196

The obtained data were fitted with different models including first-order stress decay with a 8

197

zero equilibrium stress (Eq. 6), first-order stress decay with a non-zero equilibrium stress (Eq.

198

7), Weltman model (Eq. 8) and second-order structural kinetic model (Eq. 9):

199 200 201 202 203

=

(6)

where, τ0 is the initial shear stress and k is the breakdown rate constant. −

=(

−

)

(7)

where, τeq is the equilibrium stress. =

+

(8)

204

where, A and B are the constant parameters which characterize the time-dependent behavior.

205

[(

206

where, η′0 is the initial apparent viscosity at t = 0 (i.e., structured state); η′∞ is the steady state

207

apparent viscosity at t → ∞ (i.e., non-structured state); n and k are the order of the structure

208

breakdown and the breakdown rate constant, respectively (Razavi, & Karazhiyan, 2009).

209

2.7.4. Gum viscoelastic properties

′

′ ∞ ′ ∞

)]

= ( − 1)

+1

(9)

210

Dynamic rheological measurements of aqueous dispersions (0.3%) of different BSG

211

samples were carried out at 20 °C using a controlled stress/strain rheometer (MCR 302,

212

Anton Paar) equipped with cone-plate geometry (type CP25-1). Amplitude sweep tests were

213

conducted to determine the linear viscoelastic (LVE) region at a shear strain range of 0.01%-

214

100% and constant frequency of 1 Hz. Frequency sweep tests were then carried out within

215

LVE region at a frequency range of 0.01-10 Hz and constant shear strain of 0.1%. For each

216

measurement, 2 mL of sample was carefully deposited over the rheometer plate and allowed

217

to rest for 10 min for equilibration and structure recovery before the measurement. Storage

218

modulus (G′), loss modulus (G″), loss factor (tan δ) and complex viscosity (η*) were

219

calculated from the rheological data using RheoCompass™ software (Anton Paar, Graz,

220

Austria).

9

221

2.7.5. Intrinsic viscosity

222

Dilute solution viscosity was measured using a capillary (Ubbelhode) viscometer (type

223

518 10, Schott Geräte, Hofheim, Germany) at 25 °C. Stock dispersions (5 mg/mL) were

224

prepared at room temperature and then diluted in series. The relative viscosity (ηrel) was

225

determined from Eq. 10 (Ghayour et al., 2019; Mirpoor, Hosseini, & Yousefi, 2017).

226

ηrel = tsample/tsolvent

227

where, tsample and tsolvent are the flow times of gum dispersion and solvent, respectively.

228

Specific viscosity (ηsp), reduced viscosity (ηred) and inherent viscosity (ηinh) were calculated

229

using Eqs. 11-13, respectively.

230

ηsp = ηrel - 1

(11)

231

ηred = ηsp/C

(12)

232

ηinh = lnηrel/C

(13)

233

where, C is the concentration in g/100 mL.

234

The intrinsic viscosity (ηint) was then determined using either intercept-based models

235

(Huggins (Eq. 14) and Kraemer (Eq. 15)) or slope-based models (Tanglertpaibul & Rao (Eq.

236

16), Higiro 1 (Eq. 17) and Higiro 2 (Eq. 18)) (Hosseini, et al., 2013; Naji-Tabasi, Razavi,

237

Mohebbi, & Malaekeh-Nikouei, 2016).

238

ηred = ηint + kHηint2C

(14)

239

ηinh = ηint + kKηint2C

(15)

240

)* + = 1 + [), ]-

(16)

241

)* + =

242

)* + =

243

where, ηint is the intrinsic viscosity (dL/g); kH and kK are Huggins and Kraemer constants,

244

respectively. During the intrinsic viscosity measurement, ηrel and ηsp were within the range of

245

1.2-2.0 and 0.2-1.0, respectively. Beyond this range, the effect of concentration on the

(10)

[ ./0 ]1

(17) (18)

[ ./0 ]1

10

246

viscosity is progressively increased due to the interactions of polymer chains (Qian, Cui, Wu,

247

& Goff, 2012).

248

2.7.6. Molecular conformation

249

The molecular conformation (i.e., exponent b) of BSG was estimated from the slope of

250

a double logarithmic plot of ηsp vs. C (Eq. 19) (Naji-Tabasi, Razavi, et al., 2016).

251

ηsp= aCb

252

2.8. Surface morphology

(19)

253

The morphology of freeze dried samples was characterized using a scanning electron

254

microscope (SEM, TESCAN vega3, Czech Republic) at an accelerating voltage of 20.0 kV.

255

Before characterization, samples were mounted on a holder using aluminum tape and

256

sputtered with a thin layer of gold (Desk Sputter Coater DSR1, Nanostructural Coating Co.,

257

Iran). The entropy of grayscale image was determined using Image Processing ToolboxTM for

258

MATLAB. The SEM Images were resized to 800 × 716 pixels prior to entropy measurement.

259

2.9. Interfacial properties

260

2.9.1. Interfacial tension

261

The pendant drop method using the drop shape analyzer (DSA 100, KRÜSS GmbH,

262

Hamburg, Germany) was performed to measure the ability of unmodified and modified BSG

263

to reduce the oil-water interfacial tension (IFT) (Kazemzadeh, Parsaei, & Riazi, 2015).

264

Briefly, a drop of the aqueous dispersion of hydrocolloid was formed at the tip of a needle in

265

bulk canola oil. The image of the aqueous drop was recorded using a CCD camera equipped

266

with macro lens. The IFT was determined at the leaving point of the needle tip through

267

processing the drop image, edge detection and fitting of the Laplace-Young equation.

268

2.9.2. Emulsion preparation and characterization

11

269

BSG (0.075 g) was hydrated in deionized water (17.50 g). Canola oil (7.50 g) was

270

gradually added to the aqueous phase and homogenized at 15000 rpm for 2 min using a high

271

speed homogenizer (Ultra-Turrax T18, IKA, Germany).

272

2.9.2.1. Emulsifying ability

273

The emulsifying ability of unmodified and modified BSG was investigated according to

274

the method of Chikamai, Banks, Anderson, and Weiping (1996). Briefly, 100 µL of freshly

275

prepared BSG-stabilized emulsions was diluted with 25 mL deionized water. The emulsifying

276

activity index (EAI) was calculated using the following equation.

277

EAI (m2/g)= (2 × 2.303 × D × A)/(I × Φ × C × 10,000)

278

where, A is the absorbance of diluted samples at 500 nm; I (m) is the path length of cell; D is

279

the dilution factor; Φ is the volumetric fraction of oil; C (g/mL) is the weight of hydrocolloid

280

per unit volume of the aqueous phase before emulsification; and 10,000 is the correction

281

factor for square meters.

282

2.9.2.2. Emulsion stability

(20)

283

Freshly prepared emulsions containing 0.03% sodium azide were placed in 15 mL

284

sealed tubes (height: 120 mm, internal diameter: 11 mm), and kept at ambient temperature.

285

The emulsion stability was determined over time.

286

2.9.2.3. Droplet size

287

The average droplet size of BSG-stabilized emulsions was measured over time by light

288

scattering technique (Mastersizer 2000, Malvern Instruments, Malvern, UK). Samples were

289

diluted with deionized water until an obscuration rate of 10%-20% was obtained. Mie theory

290

was applied by considering the refractive index of 1.46 and 1.33 for canola oil and water,

291

respectively. Volume-weighted mean diameter (D43) was calculated according to the Eq. 21.

12

292

D43 = ∑niDi4 ⁄ ∑niDi3

293

where, ni is the fraction of droplets corresponding to the diameter Di.

294

2.9.2.4. Zeta-potential

295

(21)

After dilution with deionized water (1:100), the zeta-potential of emulsions was

296

determined using DLS (SZ100, Horiba, Japan) at 25 °C.

297

2.9.2.5. Emulsion viscoelastic properties

298

To determine the LVE region, the amplitude sweep test was carried out at 20 °C,

299

frequency of 1 Hz and shear strain range of 0.01%-100% using an MCR 302 Anton Paar

300

rheometer (Graz, Austria). Frequency sweep test was then performed at the same temperature

301

within LVE region (shear strain 0.1%) and the frequency range of 0.01-10 Hz.

302

2.9.3. Foaming ability and foam stability

303

The ability of modified gums to stabilize the egg albumin foam was studied using the

304

method described by Naji-Tabasi and Razavi (2016). Briefly, egg albumin (0.3% w/v) was

305

added to the hydrated dispersion (20 mL) of BSG prepared at 0.3% (w/w). Whipping was

306

performed vigorously at 15000 rpm for 2 min using a high speed homogenizer (Ultra Turrax

307

T25, IKA, Germany). The head foam volume was calculated after production and after 30

308

min. Foaming ability and foam stability were calculated using the following equations.

309

Foaming ability (%) = (Vf0/V) × 100

(22)

310

Foam stability (%) = (Vf30/Vf0) × 100

(23)

311

where, Vf0, Vf30 and V are initial foam volume, the foam volume after 30 min and total

312

volume of dispersion, respectively.

313

2.10. Statistical analysis

314

All experiments were performed in at least triplicate. The results were analyzed using

315

one-way analysis of variance at the significance level of 0.05. Duncan’s multiple range tests

13

316

(SAS® software, ver. 9.1, SAS Institute Inc., NC, USA) were used to determine the

317

significant differences between the means.

318 319

3. Results and discussions

320

3.1. The extent of modification

321

The chromatograms of OSA standard solution (top panel) and OSA solution de-

322

esterified from BSG (bottom panel) are shown in Fig. 1S (supplementary data) with an

323

elution time of 4.7 min. As determined by HPLC, the amounts of esterification were 0%,

324

0.277% ± 0.09% and 1.010% ± 0.12% at OSA:BSG weight ratios (WRs) of 0, 0.01 and 0.03,

325

respectively. Shi et al. (2017) reported the esterification values of 0%, 0.64%, 1.09% and

326

1.80% at OSA to Acacia seyal gum WRs of 0, 0.01, 0.02, and 0.03, respectively. In this

327

study, the extent of BSG modification was also determined using ion mobility spectrometry

328

(IMS) likely for the first time. Proton transfer from the reactant ions to the substance is the

329

dominant ionization mechanism in corona discharge IMS (Tabrizchi, Khayamian, & Taj,

330

2000). The ionization yield of the substances is dependent on their proton affinity. The

331

chemical structure of the OSA ions developed during analysis cannot be determined without

332

performing the mass spectrometry analysis. However, the observed additional peaks in the

333

spectrum in the presence of OSA can be related to the gum modification (i.e., as a fingerprint

334

for the OSA-modified gum). The advantage of IMS over HPLC is that the alkali de-

335

esterification step of OSA from BSG backbone is not required for IMS. The IM spectra of

336

unmodified and OSA-modified BSG are shown in Fig. 2S (supplementary data). These

337

spectra were recorded in positive ion mode under the optimized conditions (section 2.3.2).

338

OSA-modified samples (denoted as S1 and S3) showed additional peaks at 8.80 ms. From

339

these spectra, the amounts of OS esterification in the samples S0, S1 and S3 were 0%, 0.31%

340

± 0.12% and 0.97% ± 0.18%, respectively. The IMS results showed a good correlation (R2 =

14

341

0.998) with those of HPLC (as the standard method) confirming the potential application of

342

IMS in determining the extent of modification.

343

3.2. Molecular weight

344

An increase in the MW was observed after modification with OSA. The average MW

345

of unmodified (C) and modified BSG samples prepared at different OSA:BSG WRs (i.e., S0,

346

S1 and S3) was 6200, 6400, 7300 and 7900 kDa, respectively. A similar increase was

347

reported by Shi et al. (2017) during the modification of Acacia seyal gum with OSA and

348

attributed to the formation of hydrophobic associations by the OSA alkyl chain. Moreover,

349

there is a possibility that both carboxyl functional groups of some OSA molecules being

350

esterified with hydroxyl groups of different biopolymer chains which leads to increase the

351

MW. Naji-Tabasi, Razavi, et al. (2016) reported that the MW of BSG fractions ranges from

352

1045 to 5980 kDa. These differences can be attributed to the differences in the source,

353

extraction method and growing conditions.

354

3.3. Zeta potential

355

Because of having carboxyl groups, BSG bears negative charge. As reported in Table 2,

356

OSA modification to a certain amount could significantly increase the negative charge of

357

BSG which was attributed to the presence of additional carboxylate groups along the BSG

358

backbone. Falkeborg and Guo (2015) similarly reported that the negative surface charge of

359

alginate was increased after modification with DSA.

360

3.4. Contact angle

361

The effect of OSA modification on the static contact angle of BSG is shown in Fig 1.

362

Since the backbone of unmodified BSG was rich in -OH groups, the added water droplet was

363

flattened more quickly, giving the lowest contact angle. The contact angle values were

364

increased after modification, indicating that OS groups were able to increase the surface

365

hydrophobicity of BSG (i.e., lower wettability by water molecules). Similarly, Chi et al.

15

366

(2007) reported that the contact angle of corn starch was gradually increased by increasing

367

the extent of hydrophobic esterification.

368

3.5. Rheological properties

369

3.5.1. Temperature effect on viscosity

370

The effect of temperature on the viscosity of BSG dispersions (1.2% w/v) is shown in

371

Fig. 2. Generally, the hydrocolloid hydration during heating increased the viscosity. Modified

372

samples showed a higher viscosity than unmodified BSG. The hydration rate was relatively

373

improved after modification. It seems that the hydration rate is a consequence of different

374

effects like changes in the gum molecular weight and surface hydrophobicity, as well as

375

distribution pattern of OS groups along BSG backbone. In spite of an increase in the

376

molecular weight and surface hydrophobicity, the possible even substitution of OS side

377

groups along BSG backbone might prevent the backbone chains from forming hydrogen

378

bonds to each other. Therefore, modified BSG was hydrated more quickly. A classic example

379

of even vs. uneven substitution is the behavior of galactomannans, like guar gum and locust

380

bean gum (LBG), in water and their interactions with xanthan gum. Guar gum is evenly

381

substituted by galactose; whereas, LBG is highly unevenly substituted. As a result, guar gum

382

is cold-water soluble, while LBG is not (Hoefler, 2004). The esterification reaction occurs

383

primarily in the amorphous regions (Wang & Wang, 2002). In spite of no significant changes

384

in the physical state after BSG modification (as obtained by XRD analysis in our previous

385

study) (Gahruie et al., 2019), a broad peak was observed at 2θ of ~29° in sample S3

386

indicating more amorphous structure, which might lead to increase the hydration rate. As

387

reported by Hosseini-Parvar, Matia-Merino, Goh, Razavi, and Mortazavi (2010), the increase

388

of viscosity with heat while shearing can also be attributed to the aggregation character of

389

BSG as a result of hydrophobic interactions, which get stronger with increasing temperature.

390

During heating at 95 °C, the decrease in the viscosity of different types of BSG at a constant

16

391

shear rate might be attributed to the thixotropic properties (section 3.5.3). An increase in the

392

viscosity was observed during cooling to 50 °C which was attributed to the higher ability of

393

biopolymer chains to organize the water molecules around themselves at lower temperatures

394

by hydrogen bonds.

395

3.5.2. Apparent viscosity

396

As shown in Fig. 3, all BSG dispersions revealed shear thinning behavior. Similar

397

observations were reported by Hosseini-Parvar et al. (2010). The shear thinning in the steady

398

shear test occurs when macromolecules are disentangled (i.e., lose their junction zones in

399

polymer solution) and become aligned in the flow direction (Behrouzian, & Razavi, 2019).

400

Shear-thinning hydrocolloids are usually utilized to modify the viscosity of foods during high

401

shear processes like filling and pumping. As reported in Table 3, the apparent viscosity of

402

unmodified (C) and modified (S0, S1 and S3) BSG dispersions (0.3% w/v) at the shear rate of

403

51.1 s-1 and 20 °C was 72.80 ±1.37, 87.25 ±21.41, 116.52 ±0.05 and 138.42 ±15.10 mPa.s,

404

respectively. Therefore, it can be stated that the use of OSA-modified BSG as a substitute for

405

native BSG increases the viscosity of final food product. A similar increase in the apparent

406

viscosity of OSA-modified Acacia seyal gum was reported by Liang, Wang, Chen, Liu, and

407

Liu (2015) and attributed to the formation of associations by hydrophobic interactions. In

408

other words, the improved thickening properties could be related to the restricted molecular

409

movements due to the intermolecular chain entanglements (Mahmood, Alamri, Abdellatif,

410

Hussain, & Qasem, 2018). Table 3 also reports the constant parameters of different

411

rheological models. Taking into account the coefficient of determination (R2) and root mean

412

square error (RMSE) values, Herschel-Bulkely and Casson models were better than Power

413

Law and Bingham models to characterize the rheological behavior of BSG and its modified

414

counterparts at the shear rate range of 1 to 62.4 s-1. The values of flow behavior index (n < 1)

415

confirmed the shear-thinning behavior and relatively decreased from 0.71 to 0.53 (in

17

416

Herschel-Bulkely model) after modification. Therefore, the OSA-modified BSG samples

417

exhibited stronger shear-thinning character which could be attributed to an increase in the

418

interaction of OSA-modified BSG with water due to the greater charge density (section 3.3)

419

and also to its higher ability to form polymers agglomerates. The larger number of

420

interactions with water results in a greater expansion of the macromolecules and the

421

formation of large 3D networks, which favors non-Newtonian behavior (Sato, Oliveira, &

422

Cunha, 2008). As discussed later (section 3.5.5), all samples revealed a random coil

423

conformation. However, the response of consistency coefficient (k) (i.e., the relative increase

424

from 0.28 to 0.83 Pa.sn) to the gum esterification could likely be related to the improved

425

stabilization induced by a higher degree of intermolecular and intramolecular interactions.

426

Similarly, higher chain–chain associations and/or entanglements between polymer chains

427

relatively increased the amount of yield stress (τ0) after modification. Naji-Tabasi and Razavi

428

(2017b) reported n, k and τ0 values of 0.36-0.48, 1.83-11.02 Pa.sn, and 0.8-3.85 Pa for

429

different fractions of 1% BSG dispersion at 20 °C, respectively. By fitting the steady shear

430

data to the Herschel-Bulkley model, Hosseini-Parvar et al. (2010) reported dynamic yield

431

stresses of 3.47 and 11.94 Pa for 1% and 2% BSG at 20 °C, respectively.

432 433

3.5.3. Time dependency

434

Various BSG dispersions revealed the thixotropic behavior over time. Shear-induced

435

network breakdown results in thixotropy, which indicates the interconnection of biopolymer

436

chains and formation of a 3D structure. Similar behavior was reported by Hosseini-Parvar

437

(2009) at a concentration range of 1% to 3% for unmodified BSG, which was more obvious

438

at higher concentration due to the stronger interactions between the polymeric chains. Table 4

439

reports the constant parameters of different models used for fitting the experimental data.

440

Taking into account R2 and RMSE values, the first-order stress decay with a non-zero

18

441

equilibrium stress was better than other models for predicting the time-dependent behavior of

442

unmodified BSG and OSA-modified BSG dispersions prepared at 0.3% concentration. The

443

breakdown rate constant (k) of sample S0 was higher than that of other BSG samples

444

indicating the presence of a network with a lower degree of interconnection, likely due to the

445

structural changes taken place in the absence of OSA during the modification process. Except

446

for the sample S0, the time-dependent behavior of BSG samples was also predictable by the

447

second order structural kinetics model. In this model, the changes in the flow behavior over

448

time are proportional to the shear-induced internal structure destruction and the breakdown

449

rate constant depends on the kinetics of the structured state/non-structured state process

450

(Abu-Jdayil, 2003).

451

3.5.4. Dynamic rheological properties

452

Many hydrocolloid dispersions are classified as viscoelastic materials and their

453

rheological properties can be investigated by dynamic rheological techniques. Several

454

mechanisms at the molecular and microscopic levels have contributions to the overall

455

material response (Deshpande, 2010). The results of strain sweep test (Fig. 4a) showed that

456

unmodified BSG and modified counterparts have a network structure or gel-like character (G'

457

> G'') in the LVE region and a liquid-like behavior (G' < G'') after the crossover (flow) point

458

(Alghooneh, & Razavi, 2019). The ability of BSG to form a relatively weak gel was similarly

459

reported by Hosseini-Parvar et al. (2010) and Rafe and Razavi (2013). The elastic and

460

viscous characters of BSG were increased after modification with OSA likely due to the

461

increase of intermolecular chain entanglements. Strong gels have linear viscoelastic

462

behaviour in higher strain and may remain in the LVE region over greater shear strains than

463

weak gels (Rafe, & Razavi, 2013). However, in this study the critical strain (i.e., the starting

464

point of gel weakening or the strain at which G′ decreases with the increase of strain) of

465

sample S3 was lower than those of other samples. This could likely be explained by the

19

466

increase in junction zones and thus the increase in the time required for new entanglements to

467

replace those disrupted by an externally imposed small deformation in the amplitude

468

oscillatory test (Alghooneh, & Razavi, 2019). Fig. 4b shows the results of frequency sweep

469

test at constant shear strain of 0.1% and frequency range of 0.01-10 Hz. In this range, all

470

BSG dispersions (0.3%) exhibited a gel-like behavior (G' > G'') either up to a critical

471

frequency (for C and S1) or over the whole experimental range (for S0 and S3). The

472

mechanical spectra showed a crossover point for samples C and S1 but not for S0 and

473

particularly S3. The crossover between G' and G'' is observed when the angular frequency (ω)

474

is equal to 1/relaxation time (trel) and indicates that the flow behavior is started (i.e., a change

475

from solid to liquid) (Deshpande, 2010; Van Vliet, 2013). Therefore, the sample C and S1

476

exhibited an elastic character at time scales larger than trel; however, the deformation of other

477

samples was elastic and recoverable. The crossover frequency in sample S1 was higher than

478

in sample C likely indicating the more rapid beginning of the elastic plateau. Modification

479

with OSA improved the viscoelastic properties of BSG due to the greater intermolecular

480

interactions and entanglements between OSA-modified BSG molecules. Tan δ (also known

481

as loss or damping factor) is the ratio of G'' to G' (Fig. 4c). Predominant elastic and viscous

482

character is observed when tan δ is <1, and >1, respectively. The sample is not a real (bulk)

483

gel when tan δ is greater than 0.1 (Mandala, Savvas, & Kostaropoulos, 2004). As shown in

484

Fig. 4c, a dominant elastic behavior (or a weak gel structure) was observed in a wide range of

485

applied frequency. Similar results were also reported by Hosseini-Parvar (2009) and Rafe,

486

Razavi, and Farhoosh (2013). Moreover, all the samples were not real gels (0.1 < tan δ), and

487

macromolecular connections were temporary and disrupted by applying high shear rates

488

(Rafe, & Razavi, 2013). The different behavior of the loss factor of sample S3 at high

489

frequency was likely due to the higher amounts of junction zones at a higher degree of

490

esterification and the dependency of the balance of bond-breaking and bond-making to the

20

491

applied frequency. An increase in the complex viscosity (η*) of BSG dispersions was

492

observed after modification with OSA (Fig. 4d), reflecting a good agreement with the results

493

of apparent viscosity (Fig. 3). Up to certain values of angular frequency, a linear decrease in

494

η* was observed and attributed to the disruption of chains entanglements and

495

macromolecules connections, confirming the shear-thinning behavior of gum dispersions.

496

The linear reduction of η* was similarly reported by Rafe and Razavi (2013); however, these

497

researchers reported the absence of a plateau (or leveling out) at a constant Newtonian value

498

over the experimental frequency range (0.01 to 10 Hz). This difference could be related to the

499

lower concentration of BSG dispersion in this study (0.3%) than in the study (1% to 3%)

500

performed by Rafe and Razavi (2013). Similar to our study, Wei et al. (2015) reported an

501

increase in η* of fenugreek gum at high frequency and ascribed to the balance between bond-

502

breaking and bond-making at different time intervals during frequency sweep test. Moreover,

503

at higher frequencies, inertia effects might play a role, while the rheometric analysis assumes

504

the absence of these (Deshpande, 2010). The slope of log η* in the linear region was -0.942, -

505

0.896, -0.862, and -0.827 for samples C, S0, S1, and S3, respectively. According to Rafe and

506

Razavi (2013), these values were steeper than the maximum value of -0.76 reported for

507

disordered polysaccharides interacting by topological entanglement. These materials are able

508

to generate weak-gel networks by fine associations of rigid and ordered molecular structures

509

in solution (Rafe and Razavi, 2013).

510 511

3.5.5. Intrinsic viscosity and molecular conformation

512

In this study, various intercept- and slope-based models (Eqs. 14-18) were used to

513

determine the intrinsic viscosity (ηint) of different BSG samples (Figs. 5a-5e). A summary is

514

reported in Table 5. To measure the ηint, the slope-based models were generally better than

515

Huggins and Kraemer models. Among which, Higiro 2 model exhibited the highest R2 and

21

516

the lowest RMSE values. The ηint values (dL/g) of unmodified BSG calculated by Huggins

517

(9.61), Kraemer (10.71), Higiro 1 (14.48) and Higiro 2 (10.70) equations were similar to

518

those reported by Mirabolhassani, Rafe, and Razavi (2016). The ηint is a unique function of

519

molecular weight and conformation. An increase in the ηint was observed after modification

520

with OSA which could be attributed to either the increase in the molecular weight (section

521

3.2) (Lapasin & Pricl, 1995) or the increase in the chain stiffness due to the increase in the

522

charge density along BSG backbone (section 3.3). The intrinsic viscosity values of

523

unmodified and modified BSG were similar to those of guar (15.80 dL/g, 25 °C), tara (14.55

524

dL/g, 25 °C), fenugreek gum (15.10 dL/g, 25 °C) (Wu, Cui, Eskin, & Goff, 2009), and LBG

525

(14.20 dL/g, 25 °C) (Sittikijyothin, Torres, & Gonçalves, 2005); but lower than other

526

hydrocolloids like κ-carrageenan (42.20 dL/g, 25 °C (Nickerson, Paulson, & Hallett, 2004)

527

and xanthan gum (110.34 dL/g, 25 °C) (Viturawong, Achayuthakan, & Suphantharika, 2008).

528

Fig. 5f shows the logarithmic plot of ηsp vs. C. The slope (exponent b) indicates the molecular

529

conformation. The values greater than 1 are associated with random coil conformation in

530

dilute regimes; whereas, those lower than unity indicate rod-like conformation (Lapasin &

531

Pricl, 1995). The exponent b of sample C, S0, S1 and S3 was 1.31, 1.31, 1.34, and 1.27,

532

respectively (Table 5); which revealed the random coil conformation even after modification.

533

The random coil conformation is a typical character of several linear polysaccharides like

534

alginate, κ-carrageenan, and LBG. The double logarithmic plot of ηsp versus C[ηint], also

535

known as master curve, is shown in Fig. 5g. Generally, the slope of the master curve in the

536

dilute and concentrated domains is 1.4 and 3.3, respectively (Mirabolhassani et al., 2016). In

537

this study, the slope of the master curve for different samples including C, S0, S1, and S3 was

538

1.44, 1.46, 1.42, and 1.41, respectively.

539

3.6. Gum microstructure

22

540

Fig. 6 shows the SEM micrographs of freeze-dried BSG samples at two different

541

magnifications (5000× and 90000×). As a measure of disorder and/or image information

542

content, the entropy of grayscale images was determined using Image Processing ToolboxTM

543

for MATLAB and amounted to 6.72, 6.30, 5.57 and, 5.54 for C, S0, S1, and S3, respectively.

544

This factor is used in the quantitative analysis and evaluation of image details. Entropy

545

measures the “complexity” of the image with respect to the spatial location of grey levels in

546

the image (Tournier, Grass, Zope, Salles, & Bertrand, 2012). In other words, this statistic is

547

high if a variable has a wide distribution over the available range and low if it has an ordered

548

and narrow distribution (Davies, 2012). Therefore, the higher value of entropy means more

549

detailed information and an image with low entropy is more homogenous than an image with

550

high entropy. In a perfectly ordered system, the entropy value is zero (Davies, 2012). The

551

decrease in entropy after modification could be related to the relative disappearance of

552

spherical particles as well as the formation of a structure with higher integrity.

553

3.7. Interfacial properties

554

3.7.1. Interfacial tension

555

The interfacial tension (IFT) between the water and oil phases in the absence of BSG

556

was 14.3 ± 0.7 mN/m. A significant decrease in the IFT was observed in the presence of BSG

557

in a manner which was dependent on the extent of modification (Table 6). The surface

558

activity of unmodified BSG was due to the presence of proteinaceous moiety (Table 1)

559

associated to the polysaccharide structure and also due to the hydrophobic character of the

560

polysaccharide itself (Osano, et al., 2014). The random coil structure of BSG also leads to the

561

surface activity (Naji-Tabasi, Razavi, et al., 2016). A significant decrease in the surface and

562

interfacial tension was similarly reported by Osano et al. (2014) depending on the gum type

563

and its concentration. Crude BSG was more effective than purified and protein-depleted BSG

564

which means that protein removal and gum purification reduce the adsorption properties of

23

565

gum to the interface. The higher ability of sample S3 to decrease the IFT was attributed to the

566

presence of larger amounts of alkyl chains along BSG backbone and hence the higher

567

hydrophobic character (section 3.4). Similar observations were reported during the

568

modification of inulin (Kokubun, Ratcliffe, & Williams, 2013) and gum Arabic (Wang et al.,

569

2014) with DSA.

570

3.7.2. Emulsifying properties

571

Emulsifying activity index (EAI) and emulsion stability refer to the ability of surface

572

active agents in the formation and stabilization of emulsions over time, respectively (Liang,

573

Wang, Chen, Liu, & Liu, 2015). The EAI of different BSG samples is shown in Fig. 7. The

574

EAI of sample C was lower than that of sample S0. Because of non-significant differences in

575

the gum chemical composition after modification with OSA (Table 1), the higher activity of

576

S0 than C might be attributed to the “gum maturation” during the treatments applied in the

577

absence of OSA. Maturation is a process in which the associations of arabinogalactan (or

578

other polysaccharide molecules) and low MW glycoproteins are formed (Al-Assaf, Phillips,

579

Aoki, & Sasaki, 2007). In agreement with the results of IFT, BSG modification improved the

580

EAI. A higher activity was observed at higher OSA:BSG WR. A favorable balance in the

581

hydrophilic-hydrophobic character of modified BSG resulted in improving the ability of

582

hydrocolloid to form O/W emulsions.

583

Table 7 reports the physical stability of emulsions during storage. Generally, all

584

emulsions revealed appropriate stability mainly due to the inherent surface activity of

585

unmodified BSG and its improvement in the modified counterparts (Table 6). Hydrocolloids

586

also contribute to the stability of emulsions through increasing the viscosity of continuous

587

phase (Dokić, Dokić, Dapčević, & Krstonošić, 2008; Xu, Huang, Fu, & Jane, 2015). In this

588

study, the OSA-modified BSG exhibited a higher ability than unmodified BSG in increasing

589

the viscosity.

24

590

The separation of continuous phase was started after 7 d in the emulsions stabilized

591

with BSG C and BSG S0 (Fig. 8 and Table 7) and reached to the final stability of 85.5% and

592

93.25% after 42 d, respectively. The emulsions prepared by sample S3 remained completely

593

stable over the entire storage period. It should be noted that the emulsification method has a

594

vital role in the final stability. As mentioned already, Osano et al. (2014) reported that a two-

595

stage high pressure homogenization led to the formation of stable emulsions with monomodal

596

droplet distribution and average droplet size (d32) below 1 µm at 0.3% BSG concentration.

597

Utilization of an Ultraturrax to develop the O/W emulsions, as performed in the current

598

study, normally leads to larger droplet size. Therefore, lower stability in the control samples

599

was observed as compared to the emulsions prepared and evaluated by Osano et al. (2014). A

600

significantly higher stability was observed in the samples emulsified with the OSA-modified

601

BSG. The increase in the stability was attributed to the smaller droplet size and improved

602

viscoelastic properties.

603

Fig. 9 (a-c) shows the droplet size distribution of different BSG-stabilized emulsions

604

over 42-d storage. A bimodal droplet distribution was observed in the emulsions stabilized

605

with samples C, S0, and S1. A shift to the left side (i.e., a decrease in the average droplet

606

size) was noted in the emulsions stabilized with S1 (Fig. 10a). A monomodal droplet

607

distribution and significantly smaller average droplet size were observed in the emulsion

608

stabilized by sample S3 at the first day of storage (Fig. 9a); which was attributed to the higher

609

hydrophobicity of S3 (Fig. 1) and its higher kinetics of adsorption to the oil-water interface

610

(Wang et al., 2014). Changes in the droplet size distribution were dependent on the BSG type

611

(Fig. 9). Generally, an increase in the average droplet size was observed over time (Fig. 10a),

612

which was attributed to the droplet coalescence and flocculation. The least increasing rate in

613

the average droplet size was observed in the S3-stabilized emulsion.

25

614

In addition to the smaller droplet size, the higher physical stability of the emulsion

615

prepared with S3 could also be attributed to its superior elastic character. As shown in the

616

results of strain sweep test (Fig. 11a), O/W emulsions stabilized with unmodified and

617

modified BSG have gel-like character (G' > G'') in the LVE region and a liquid-like character

618

(G' < G'') after the crossover point. The nonlinear response could be related to the large

619

deformations, structural changes, and phase transitions (Deshpande, 2010). The viscoelastic

620

characters of emulsions were improved after modification with OSA. The emulsion rheology

621

is strongly affected by the state of flocculation (Guerrero, Partal, & Gallegos, 1998).

622

Unflocculated or weakly flocculated emulsions show a crossover point between G' and G''

623

(Deshpande, 2010). The emulsion stabilized with sample S3 remained in the LVE over

624

greater shear strains which indicated a higher degree of interactions between the emulsifiers

625

from adjacent droplets. Osano et al. (2014) similarly reported that the BSG-stabilized

626

emulsions had a gel-like character at all concentrations. The results of frequency sweep test

627

and loss factor (Fig. 11b and 11c) showed that BSG and its modified counterparts were able

628

to stabilize O/W emulsions through developing a weak gel structure. In agreement with the

629

results of gum viscoelastic properties (section 3.5.4), the storage and loss moduli of BSG-

630

stabilized emulsions were increased after modification. The emulsion gel systems were not

631

real (0.1 < tan δ) and the connections could be disrupted at high shear rates. The shear-

632

thinning behavior of emulsions (Fig. 11d) was attributed to the shear-induced deflocculation

633

of oil droplets (Boostani et al., 2019).

634

Fig. 10b shows the zeta potential of different BSG-stabilized O/W emulsions as a

635

function of time. The negative value of zeta potential was due to the negative charge of BSG

636

molecules. A decrease in the zeta-potential of BSG-stabilized emulsions by increasing the

637

acidity and ionic strength was reported by Hosseini-Parvar et al. (2016). In addition to the

638

smaller droplet size and improved viscoelastic properties, the strong electrostatic repulsion

26

639

between the dispersed droplets could contribute to the emulsion stability. The steric repulsion

640

is likely the main mechanism behind the stability of BSG-stabilized emulsions (Jayme,

641

Dunstan, & Gee, 1999). Higher MW and amphiphilic character generally improve the steric

642

stabilization (Jayme, et al., 1999; Xu, et al., 2015). Li, Fu, Luo, and Huang (2013) reported

643

that the higher MW of esterified starch was beneficial in the stabilization of emulsions.

644 645

3.7.3. Foaming properties

646

Table 8 reports the effect of BSG modification on the foaming ability of egg albumin

647

and the final foam stability. A significant increase (from 41.67% to 59.09%) in the foaming

648

ability was observed after BSG modification with OSA. Naji-Tabasi and Razavi (2016)

649

reported that the foaming capacity of 0.3% BSG dispersion (containing 0.3% albumin) was

650

28%. The ability of different BSG fractions in increasing the foaming ability was attributed to

651

their surface activity and MW (Naji-Tabasi, and Razavi, 2016). The surface activity of gums

652

is rooted in the molecular characteristics like the hydrophobic character of side groups

653

attached to the hydrophilic backbone or the presence of a proteinaceous component linked

654

covalently or physically to the polysaccharide (Dickinson, 2009). In this study, the increase in

655

the foaming capacity was attributed to the additive effect of OSA-modified BSG. In fact, the

656

presence of hydrophobic OS groups along the BSG backbone resulted in increasing the

657

surface activity of BSG and hence improving the formation of liquid film at the surface of gas

658

bubbles (Amid, Mirhosseini, Poorazarang, & Mortazavi, 2013). Without having appropriate

659

hydrophobic characteristics, high MW polysaccharides do not have a tendency to adsorb at

660

air-water interface; however, they are likely able to improve the protein foam stability via

661

working as a thickener or a gelling agent (Koocheki, Razavi, & Hesarinejad, 2012). Foam

662

stability is the ability of foam to maintain some characteristics (e.g., bubble size, foam

663

volume and liquid content) constant over time (Naji, Razavi, & Karazhiyan, 2012).

27

664

Generally, the gas bubbles rise to the top and undergo deformation (Jahanbin, Moini, Gohari,

665

Emam-Djomeh, & Masi, 2012). An increase in the albumin foam stability was observed by

666

increasing the extent of BSG modification (Table 8). The foam stability is dependent on the

667

aqueous phase viscosity (Koocheki, Taherian, & Bostan, 2013). Therefore, the OSA-induced

668

surface activity and the higher viscosity of modified gums in the aqueous phase were mainly

669

responsible for increasing the foam stability. The network formation, as a result of higher

670

viscosity, reduces the coalescence of air-bubbles (i.e., disproportionation) and hence

671

increases the foam stability (Jahanbin et al., 2012). Moreover, the film permeability is

672

influenced by the presence of polysaccharide molecules and their chemical modification.

673

Makri and Doxastakis (2007) reported that LBG, gum Arabic, xanthan gum and a mixture of

674

xanthan/LBG have the ability to improve the foam stability due to the network formation.

675

Asghari, Norton, Mills, Sadd, and Spyropoulos (2016) reported that the addition of OSA-

676

modified starch to protein system significantly increased the foam capacity.

677 678

4. Conclusion

679

Basil seed gum was modified using various amounts OSA. The extent of esterification

680

was determined using HPLC and IMS. OSA modification significantly improved the

681

rheological and interfacial properties of BSG. An increase in the MW, contact angle, zeta-

682

potential, intrinsic viscosity, apparent viscosity, and storage modulus was observed after

683

modification. A higher emulsion and foam stability was observed at a higher degree of

684

modification. The results of this study showed that the OSA-modified BSG is a good

685

candidate for utilization as a thickener and emulsifier in food products (e.g., dressings) which

686

need long-term stability.

687 688

Acknowledgment 28

This work was financially supported by Shiraz University (Grant number

689 690

96GCU5M194065).

691 692

Conflict of interest

693

None

694

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34

Table 1. Chemical composition of unmodified and modified BSG samples Sample

C

S0

S1

S3

Moisture

5.74 ± 0.10 a

5.67 ± 0.08 a

5.68 ± 0.05 a

5.70 ± 0.13 a

Protein

1.28 ± 0.05 a

1.21 ± 0.02 a

1.23 ± 0.02 a

1.23 ± 0.03 a

85.09 ± 0.46 a

85.02 ± 0.67 a

84.87 ± 0.82 a

Carbohydrate 84.74 ± 0.86 a

Data are the average of three independent replicates ± standard deviation. C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively. In each row, different lowercase letters indicate significant differences (p<0.05).

52

Table 2. Effect of OSA modification on the zeta-potential of BSG Sample

C

S0

S1

S3

Zeta potential (mV)

-57.83±1.18 b

-59.83±1.19 b

-59.50±0.36 b

-66.07± 1.27 a

Data are the average of three independent replicates ± standard deviation. C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively. Different lowercase letters indicate significant differences (p<0.05).

53

Table 3. Apparent viscosity and rheological parameters of different BSG dispersions (0.3%) at 20 °C Sample Apparent viscosity (mPa.s) Model Constant Power Law k (Pa.sn) n R2 (%) RMSE

C S0 S1 S3 72.80±1.37 c 87.25±21.41 bc 116.52±0.05 ab 138.42±15.10 a 0.80±0.19 b 0.48±0.02 a 93.16±8.78 a 0.13±0.12 a

0.89±0.34 ab 0.40±0.03 ab 94.86±2.40 a 0.11±0.04 a

1.26±0.23 ab 0.40±0.05 ab 97.33±0.50 a 0.08±0.02 a

1.75±0.47 a 0.36±0.04 b 98.24±0.40 a 0.06±0.01 a

Casson

k (Pa.s0.5) 0.20±0.01 a 0.81±0.03 b τ0 (Pa) R2 (%) 93.70±6.46 a RMSE 0.09±0.06 a

0.19±0.01 a 0.90±0.11 ab 98.42±0.24 a 0.04±0.01 a

0.22±0.02 a 1.0±0.06 ab 98.50±0.02a 0.05±0.00 a

0.22±0.00 a 1.10±0.09 a 98.65±0.96 a 0.05±0.02 a

Bingham

µ (Pa.s) τ0 (Pa) R2 (%) RMSE

0.07±0.01 b 1.19±0.46 b 97.74±0.35a 0.57±0.55 a

0.09±0.01 ab 1.75±0.25 ab 96.02±0.30 a 0.36±0.03 a

0.10±0.00 a 2.38±0.63 a 95.13±1.94 a 0.66±0.50 a

0.07±0.01 b 0.87±0.10 b 91.58±5.54 a 0.43±0.32 a

k (Pa.sn) 0.28±0.22 a 0.21±0.05 a 0.53±0.10 a 0.83±0.37 a n 0.71±0.25 a 0.73±0.02 a 0.59±0.02 a 0.53±0.09 a τ0 (Pa) 0.51±0.46 a 0.85±0.36 a 0.87±0.39 a 1.12±0.15 a R2 (%) 98.89±1.36 a 98.88±0.76 a 99.52±0.15 a 99.80±0.12 a RMSE 0.16±0.08 a 0.12±0.04 a 0.15±0.02 a 0.14±0.05 a Data are the average of three independent replicates ± standard deviation. C indicates unmodified sample.

Herschel-Bulkely

S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively. In each row, different lowercase letters indicate significant differences (p<0.05).

54

Table 4. Time-dependent characteristics of BSG dispersions (0.3%) obtained by different models First-order stress decay with a zero equilibrium stress model τ0

C 101.31± 0.76d S 116.48± 0 4.16c S 143.26± 1 0.38b S 187.89± 3 6.30a

First-order stress decay with a non-zero equilibrium stress model

Second-order structural kinetic model

Weltman model

k

R2

RMSE

τ0

τeq

k

R2

RMSE

B

A

R2

RMSE

η0/η

k

R2

RMSE

2.60± 0.71a 1.70± 1.45a 3.18± 2.41a 4.63± 0.66a

0.66± 0.03a 0.55± 0.30a 0.49± 0.18a 0.78± 0.20a

1.45± 0.29a 0.97± 0.33a 2.31± 1.08a 1.82± 0.96a

52.33±2 .65b 99.14±8 .29a 96.28±3 .85a 105.00± 4.94a

44.85± 1.51d 54.96± 0.42c 61.39± 1.58b 80.60± 1.78a

0.09± 0.04b 0.30± 0.03a 0.13± 0.02b 0.07± 0.01b

0.93± 0.01a 0.94± 0.01a 0.98± 0.02a 0.95± 0.01a

0.29± 0.03ab 0.21± 0.13b 0.30± 0.07ab 0.50± 0.08a

0.0003± 0.0001a 0.0002± 0.0001a 0.0002± 0.0002a 0.0003±0 .0001a

92.39± 3.09d 110.81± 0.40c 132.02± 7.69b 172.67± 5.04a

0.33± 00.4a 0.31± 0.28a 0.24± 0.08a 0.52± 0.28a

0.02± 0.00a 0.01± 0.00a 0.02± 0.01a 0.02± 0.00a

2.12± 0.06b 1.40± 0.07c 2.97± 0.06a 2.31± 0.12b

0.99± 0.01a 0.98± 0.02a 0.99± 0.01a 0.98± 0.01a

0.92± 0.01a 0.76± 0.09b 0.93± 0.01a 0.93± 0.01a

0.46± 0.14ab 0.36± 0.13b 0.70± 0.02ab 0.75± 0.16a

Data are the average of three independent replicates ± standard deviation. C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively. In each column, different lowercase letters indicate significant differences (p<0.05).

55

Table 5. Molecular conformation parameters and intrinsic viscosity values (dL/g) of different BSG samples obtained by various intercept- and slope-based models Huggins ηint C 9.61± 1.03a S 9.18± 0 2.01a S 11.32± 1 1.32a S 12.24± 3 2.06a

RMSE R2 0.97± 1.13± 0.02a 0.31a 0.98± 1.44± 0.01a 0.96a 0.99± 1.00± 0.01a 0.62a 0.96± 1.67± 0.04a 0.86a

Kraemer ηint 10.71± 0.99a 10.41± 1.94a 12.39± 1.23a 13.40± 1.87a

RMSE R2 0.92± 0.77± 0.09a 0.36a 0.93± 1.09± 0.02a 0.98a 0.97± 0.68± 0.01a 0.67a 0.79± 1.14± 0.28a 0.90a

Tanglertpaibul & Rao ηint 20.59± 0.40c 21.08± 0.71bc 22.64± 0.56ab 23.70± 0.75a

RMSE R2 0.94± 0.10± 0.00a 0.00a 0.94± 0.11± 0.01a 0.01a 0.95± 0.10± 0.01a 0.00a 0.95± 0.11± 0.01a 0.00a

Higiro 1 ηint 14.48± 0.22c 14.72± 0.41bc 15.56± 0.31ab 16.07± 0.42a

RMSE R2 0.99± 0.03± 0.01a 0.00a 0.99± 0.04± 0.01a 0.01a 0.99± 0.03± 0.00a 0.01a 0.99± 0.03± 0.00a 0.01a

Higiro 2 ηint 10.70± 0.13c 10.81± 0.25bc 11.30± 0.18ab 11.55± 0.25a

RMSE R2 0.99± 0.01± 0.00a 0.00a 0.99± 0.01± 0.00a 0.00a 0.99± 0.02± 0.00a 0.01a 0.99± 0.02± 0.00a 0.01a

Molecular conformation a b 51.43± 1.31± 6.79a 0.05a 55.59± 1.31± 8.58a 0.04a 60.25± 1.34± 12.64a 0.09a 52.99± 1.27± 8.33a 0.06a

Data are the average of three independent replicates ± standard deviation. C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively. In each column, different lowercase letters indicate significant differences (p<0.05).

56

Table 6. Effect of OSA modification on the interfacial tension (IFT) of different BSG samples Sample

C

S0

S1

S3

IFT (mN/m) 10.68±0.83 a 7.72±0.51 b 7.51±0.30 b 3.06± 0.42 c

Data are the average of three independent replicates ± standard deviation. C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively. Different lowercase letters indicate significant differences (p<0.05).

57

Table 7. Physical stability of BSG-stabilized O/W emulsions during storage Storage time (day) Sample

0

1

7

21

42

C

100.00±0.00 Aa

100.00±0.00 Aa

90.70±0.42 Cb

88.75±0.35 Cc

85.50±0.71 Dd

S0

100.00±0.00 Aa

100.00±0.00 Aa

97.15±1.20 Bab

95.30±1.84 Bbc

93.25±1.06 Cc

S1

100.00±0.00 Aa

100.00±0.00 Aa

100.00±0.00 Aa

100.00±0.00 Aa

97.70±0.42 Bb

S3

100.00±0.00 Aa

100.00±0.00 Aa

100.00±0.00 Aa

100.00±0.00 Aa

100.00±0.00 Aa

Data are the average of three independent replicates ± standard deviation. C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively. Different capital letters (in each column) and different small letters (in each row) indicate significant differences (p<0.05).

58

Table 8. Foaming ability and foam stability of egg albumen foam in the presence of different types of BSG Sample Foaming ability (%) Foam stability (%) C

41.67±1.31c

69.10±2.84bc

S0

42.42±1.31c

67.93±4.55c

S1

53.03±2.62b

75.88±5.55ab

S3

59.09±4.55a

77.03±2.09a

Data are the average of three independent replicates ± standard deviation. C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratios of 0, 0.01 and 0.03, respectively. In each column,

different

lowercase

significant differences (p<0.05).

59

letters

indicate

Fig. 1. Effect of modification on the contact angle of freeze-dried BSG samples; C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively. Different letters indicate significant differences (p<0.05).

35

600

C

S0

S1

S3

100

Temperature (°C)

550

90 80

450 400

70

350

60

Temperature (°C)

Viscosity (cP)

500

300 50

250 200

40 0

100

200

300

400

500

600

700

800

Time (s) Fig. 2. Effect of temperature on the viscosity of different types of BSG obtained by Rapid Visco Analyser; C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively.

36

Apparent viscosity (mPa.s)

10000

1000

100

10 C

S0

S1

S3

1 1

10 Shear rate (s-1)

100

Fig. 3. Changes in the apparent viscosity of different BSG dispersions (0.3%) at 20 °C as a function of shear rate. C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively.

37

G′ and G″ (Pa)

100

(a)

C, G′

S0, G′

S1, G′

S3, G′

C, G″

S0, G″

S1, G″

S3, G″

10

1 0.01

1000

0.1

(b)

1 Shear Strain (%)

10

100

C, G′

S0, G′

S1, G′

S3, G′

C, G″

S0, G″

S1, G″

S3, G″

G′ and G″ (Pa)

100

10

1

0.1 0.01

0.1

1 Angular Frequency (rad/s)

38

10

100

Loss factor (tan δ)

10

(c)

S0

S1

S3

1 Angular Frequency (rad/s)

10

1

0.1 0.01

1000

Complex viscosity, η* (Pa.s)

C

0.1

100

(d)

100

C

S0

S1

S3

10

1

0.1 0.01

0.1

1 Angular Frequency (rad/s)

39

10

100

Fig. 4. (a) and (b): Amplitude and frequency sweep test of different BSG dispersions (0.3%) performed at 20 °C, respectively; (c) and (d): changes in the in loss factor and complex viscosity of BSG dispersions as a function of angular frequency, respectively. C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively.

40

35

19

(a: Huggins)

(b: Kraemer)

Inherent viscosity (ηinh)

Reduced viscosity (ηred)

18 30 25 20 15

17 16 15 14 13 12 11

10 0

2.4

0.02 0.04 Concentration (g/dL)

0

0.06

0.9

(c:Tanglertpaibul & Rao)

Relative viscosity (ηrel)

0.06

(d: Higiro 1)

0.8

2.2

0.7 2

0.6 Lnηrel

1.8 1.6

0.5 0.4 0.3

1.4

0.2

1.2

0.1

1

0 0

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0

0.02 0.04 Concentration (g/dL)

0.06

0

0.02 0.04 Concentration (g/dL)

0.06

0.2

(e: Higiro 2)

(f)

0 -0.2 Log ηsp

ηrel-1

0.02 0.04 Concentration (g/dL)

-0.4 -0.6 -0.8 -1

0

0.02 0.04 Concentration (g/dL)

-1.2

0.06

-2.4

41

-2 -1.6 Log C (g/dL)

-1.2

1

(g: master curve)

Log ηsp

0.5 0

-0.5 -1 -1.5 -1.5

-1

-0.5 Log C[ηint]

0

0.5

Fig. 5. (a)-(e): the plots of various intercept-based (a-b) and slope-based (c-e) models for measuring the intrinsic viscosity (ηint) of different BSG samples; (f): double logarithmic plot of specific viscosity (ηsp) vs. concentration; (g): master curve of different samples; C (

) indicates unmodified

sample. S0 ( ), S1 ( ), and S3 ( ) indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively.

42

Fig. 6. SEM micrographs of freeze-dried BSG at two magnifications (5000 × (top panel) and 90000 × (bottom panel)); C indicates unmodified sample. S0, S1 and S3 indicate the modified samples prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively.

43

45

a b

EAI (m2/g)

40 c

35 d 30 25 C

S0

S1

S3

Sample Fig. 7. Emulsifying activity index (EAI) of different types of BSG; C indicates the emulsion stabilized with unmodified BSG. S0, S1 and S3 indicate those stabilized with the modified gums prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively. Different lowercase letters indicate significant differences (p<0.05).

44

Fig. 8. Visual observation of continuous phase separation (red arrows) in different BSG-stabilized O/W emulsions during storage at ambient temperature; C indicates the emulsion stabilized with unmodified BSG. S0, S1 and S3 indicate those stabilized with the modified gums prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively.

45

7

(a)

Volume (%)

6

C

S0

S1

S3

5 4 3 2 1 0 0.01

8 7

0.1

1

10 100 Particle size (µm)

1000

10000

1000

10000

1000

10000

(b) C

S0

S1

S3

Volume (%)

6 5 4 3 2 1 0 0.01

10

Volume (%)

8

0.1

1

10 100 Particle size (µm)

(c) C

S0

S1

S3

6 4 2 0 0.01

0.1

1

10 100 Particle size (µm)

46

Fig. 9. Droplet size distribution of different BSG-stabilized O/W emulsions over time (a: 1st day; b: 7th day; and c: 42nd day); C indicates the emulsion stabilized with unmodified BSG. S0, S1 and S3 indicate those stabilized with the modified gums prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively.

47

140

(a)

C

S0

120

S1

S3

Aa

Ab

Aa

D43 (µm)

100 80

Ac

Ab Ab Ba

60 Bb

Bb

40 20

Ca

Cb

Cb

0 1

7 Storage time (d)

(b)

42

Storage time (d) 1

7

14

42

AbAbAbcAa

Aa ABa BCb Ca

0

Zeta potential (mV)

-10 -20 -30 -40 -50 Ba -60 -70

CabCa

Ac Aa

Ab

Aa

Bc C

S0

S1

S3

Fig. 10. Changes in the (a) average droplets size and (b) zeta potential of different BSG-stabilized emulsions during storage at room temperature; C indicates the emulsion stabilized with unmodified BSG. S0, S1 and S3 indicate those stabilized with the modified gums prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively. At a same time of storage, different capital letters indicate significant (p<0.05) differences between different emulsions. For a same type of emulsion, different lowercase letters indicate significant (p<0.05) differences over time.

48

100

(a)

G′ and G″ (Pa)

10

1

0.1 0.01

C, G′

S0, G′

S1, G′

S3, G′

C, G″

S0, G″

S1, G″

S3, G″

0.1

1 Shear Strain (%)

100

10

100

(b)

1000

G′ and G″ (Pa)

10

100

C, G′

S0, G′

S1, G′

S3, G′

C, G″

S0, G″

S1, G″

S3, G″

10

1 0.01

0.1

1 Angular Frequency (rad/s)

49

10

(c)

Loss Factor

C

S0

S1

S3

1

0.1 0.01

0.1

Complex viscosity (Pa.s)

1000

1 Angular Frequency (rad/s)

10

100

(d) C

S0

S1

S3

100

10

1 0.01

0.1

1 Angular Frequency (rad/s)

50

10

100

Fig. 11. (a) and (b): amplitude and frequency sweep test of freshly prepared BSG-stabilized O/W emulsions performed at 20 °C, respectively; (c) and (d): changes in the in loss factor and complex viscosity as a function of angular frequency, respectively. C indicates the emulsion stabilized with unmodified BSG. S0, S1 and S3 indicate those stabilized with the modified gums prepared at OSA:BSG weight ratio of 0, 0.01 and 0.03, respectively.

51

Highlights: •

Basil seed gum (BSG) was modified using OSA at various OSA contents.

•

The extent of modification was measured using HPLC and ion mobility spectrometry.

•

Modification induced an increase in viscosity, molecular weight and zeta-potential.

•

Modification improved the emulsifying and foaming properties of BSG.

•

Modification increased the storage modules of BSG and BSG-stabilized emulsions.

Declarations of interest: none