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Bulk, foam and interfacial properties of tannic acid/sodium caseinate nanocomplexes Fuchao Zhan, Jing Li, Yuntao Wang, Minqi Shi, Bin Li, and Feng Sheng J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.8b00503 • Publication Date (Web): 08 Jun 2018 Downloaded from http://pubs.acs.org on June 8, 2018

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Journal of Agricultural and Food Chemistry

1

Bulk, foam and interfacial properties of tannic acid/sodium caseinate

2

nanocomplexes

3

Fuchao Zhan†, §, Jing Li†, §, Yuntao Wang∥, Minqi Shi†, Bin Li*,†, § , and

4

Feng Sheng*, ‡

5 6 7

†

College of Food Science and Technology, Huazhong Agricultural

University, Wuhan 430070, China ‡

Hubei Collaborative Innovation Center for Green Transformation of

8

Bio-Resources, The College of Life Sciences, Hubei University, Wuhan,

9

430062, China

10

§

Key Laboratory of Environment Correlative Dietology (Huazhong

11

Agricultural University), Ministry of Education, Wuhan 430070, China

12

∥

13

Light Industry, Zhengzhou 450003, China

14

*

School of Food and Biological Engineering, Zhengzhou University of

Corresponding authors:

15

Bin Li, Email address: [email protected]

16

Feng Sheng, Email address: [email protected]

17 18 19 20 21 22

ACS Paragon Plus Environment


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ABSTRACT: For this work, the aim was to investigate the adsorption of

24

the tannic acid (TA)/sodium caseinate (SC) nanocomplexes at the

25

air/water interface，then to research its relationship with foam properties.

26

Firstly, nanocomplexes was prepared in a different mass ratio of TA and

27

SC. The bulk behavior of nanocomplexes was evaluated by Dynamic

28

light scattering (DLS), signal-intensifying fluorescence probe (ANS) etc.

29

As the concentration of TA increased, the z-Average Diameter (Dz) of

30

TA/SC nanocomplexes decreased gradually and the negative charge

31

increased. Meanwhile, the surface hydrophobicity(So) of the SC also

32

decreased after the addition of TA. The interfacial properties were

33

determined by dynamic surface tension and dilational rheology. The

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presence of polyphenols decreased the surface pressure (π) that resulted

35

in poor foamability. However, the elastic (Ed) component of the dilational

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modulus of films also increased as polyphenols concentration increased,

37

which gave rise to admirable foam stability. The contribution of

38

polyphenols to stabilize foam columns may be caused by interfacial

39

interaction between proteins and polyphenols.

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KEYWORDS: Tannic acid; Sodium caseinate; Interfacial properties;

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Foam; Stability

42 43 44


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INTRODUCTION

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Many common foods in daily life such as marshmallow, beer, mousse,

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meringue, nougat, ice cream and wine are composed of foam and other

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multi-phase food system1-3. The adsorption kinetics and the dilational

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rheology of surface-active agent are the main factors that determine the

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formation and stability of multi-phase foamed systems4-9. As a polymeric

51

surfactant with multiple anchoring sites at the interface, protein can be

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used to stabilize the interface layer through the unfolding of adsorbed

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protein molecules. This behavior has an important contribution to the

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interfacial rheological properties, and the protein is attached to the

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adsorption layer10, 11. However, for the formation of stable protein foam ,

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excellent surface properties are required, such as the rapid diffusion of

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protein molecules from the bulk phase to the interfacial phase and the

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marvelous

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development of technology and method to improve the functional

60

characteristics of protein is in great demand for foaming agents.

ability to

reduce interfacial pressure

12

.

Therefore,

61

The interactions between proteins and polyphenols have strong

62

influence on the stability of protein-based colloidal systems, which have

63

been continuously studied by many researchers. For example, the

64

combination of polyphenol compounds and pea protein might improve

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the antioxidant capacity of pea protein during heating13. The presence of

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polyphenols can impact the capacity of the proteins to interact at the



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air/water or oil/water interface via inducing cross-linking of the adsorbed

68

proteins14, 15. In addition to such desirable effects, the interaction of

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protein-polyphenols may also cause adverse impact such as undesirable

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haze in beer16, wine and clear fruit juices17, 18 and astringency of various

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beverages19, 20, which is attributed to insoluble complexes resulted from

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interactions between proteins and polyphenols.

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Tannic acid (TA) is a kind of water-soluble polyphenol compounds

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which can be found in many other types of plants. This polyphenol with

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large molecular weight contain abundant catechol, and pyrogallol thereby

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can interact with biological macromolecules21, 22. According to previous

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report, the combination of tannic acid and gelatin has positive influence

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on antioxidant capacity and emulsion stability of fish oil-in-water

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emulsion23. Moreover, TA also has many other kinds of extraordinary

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biological activities, such as hemostatic, antibacterial properties and

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antitumor progression24-26. In some recent studies, it was found that the

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protein-polyphenol mixtures display remarkable influence on adsorption

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behavior of protein at the interface. Therefore, understanding and

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regulating the complex surface behavior of protein-polyphenol mixtures

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is extremely necessary, which is a crucial issue for the formation and

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stability of food systems.

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In this work, the major aims are (1) to characterize the interaction

88

between protein and polyphenol in bulk phase; (2) to study the dilational


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rheological properties of tannic acid/sodium caseinate nanocomplexes at

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the air/water interface; (3) to investigate and compare the relationship

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between interfacial behavior and the foam properties of TA/SC systems.

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In order to realize these objectives, the TA/SC interactions in bulk were

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first investigated by using the Dynamic light scattering (DLS) and laser

94

Doppler velocimetry(LDV) measurements. Sodium caseinate(SC), the

95

main component of milk proteins, is used in this research due to its

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comprehensively

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hydrophobicity of the mixed TA/SC systems are detected. surface

98

rheological measurements were performed to study the dynamic surface

99

tension and interface dilational rheological of tannic acid/sodium

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caseinate system. Finally, the foam properties of TA/SC nanocomplexes

101

were evaluated. The correlation between the interactions in bulk,

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interfacial behaviors and corresponding functional properties of the

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TA/SC

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stabilization mechanism was proposed to elucidate the influence of

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protein structure modification on the interface properties.

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MATERIALS AND METHODS

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Materials. Tannic acid was obtained from Aladdin Chemical Co., China.

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Sodium caseinate from bovine milk, 1-anilinonaphthalene-8-sulfonic

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acid(ANS) and Folin-Ciocalteau reagent used in this study were bought

110

from Sigma-Aldrich (St. Louis, MO). All other chemicals used were of

were

studied

confirmed,

properties.

Subsequently,

simultaneously,

a


possible

the

surface

synergistic


111

analytical grade, and Milli-Q purified water was used in all experiments.

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Preparation of the polyphenol-protein nanocomplexes. Tannic acid

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(TA) and sodium caseinate (SC) powders were dissolved in phosphate

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buffer solution (pH 6.0, 0.01M) at 25 ℃. HCl (0.01 N) or NaOH (0.01N)

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was used for pH adjustment. The TA/SC nanocomplexes were prepared

116

by mixing appropriate volumes of each solution. Then, the mixed systems

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were stored overnight at 4 ℃.

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z-Average Diameter (Dz) and Zeta Potential measurements. The

119

particle size distributions and z-Average Diameter (Dz) of the samples

120

were determined by using Zetasizer Nano-ZS instrument (Malvern

121

Instruments Ltd, UK). The average hydrodynamic diameter of the

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particles in bulk was calculated based on Stokes-Einstein equation27.

123

The zeta potential of each sample was determined using the Zetasizer

124

Nano-ZS instrument (Malvern Instrument Ltd., UK) with laser Doppler

125

velocimetry technique. The diluted sample was loaded in the cell, then a

126

voltage was applied. The measurements were conduct at 25 ℃.

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Samples with single components (Sodium caseinate or Tannic acid)

128

were filtered with 0.45 or 0.22 µm filters prior to analysis and use. TA

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solution was added into SC solution until the final mass ratio of TA/SC

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up to 0, 0.1, 0.3, 0.5 and 1, respectively. Meanwhile, the particle size and

131

zeta potential were measured after dilution of protein concentration to

132

0.1%(w/v). The analyses were carried out in 3 repetitions.


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Determination of total phenolics and percentage combined to SC. The

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Folin-Ciocalteu spectrophotometric method28 was used to determine the

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total phenolic and the polyphenol combined with SC. Briefly, a gallic

136

acid stock solution (1mg/mL) was used to prepare a calibration standard

137

curve. The tannic acid (10 ml) was diluted to 50 ml with Milli-Q purified

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water. The TA solution was diluted to 1mg/mL for the purpose of

139

preparing standard curve (Absorbance values between 0.2 and 0.7). Then,

140

the 300 µL Folin-Ciocalteau reagent and 400 µL 10% Na2CO3 solution

141

were added to TA solution (1mg/mL, 50µL), respectively. The contents

142

were diluted with water to 5 mL, heated for 1 h at 30℃. Then, the

143

absorbance of the green color was determined at 765 nm with UV-Vis

144

spectrophotometer(UV-1100, MAPDA). The concentration of phenolics

145

was determined by comparison with the standard curve of gallic acid. The

146

analyses were carried out in 3 repetitions.

147

The TA/SC mixtures were ultrafiltrated for 15 min at 4000 rpm to get

148

the free polyphenols (ultrafiltration with cutoff 10kDa). Besides, the

149

protein in the filtrate alone was determined to avoid interference for the

150

absorbance. The percentage of TA combined with SC was evaluated as

151

the following equation29: polyphenols boundሺ%ሻ=

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Total polyphenol-polyphenol in filtrated ×100% Total polyphenol


(1)


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mg of polyphenol s bound per mg of protein Total polyphenol (mg) × Polyphenols bound(%) = mg of protein × 100

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(2)

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Surface hydrophobicity (So). Change in So of SC samples after TA

155

treatments were determined according to methods described in previous

156

research30, 31 with ANS as a fluorescence probe. TA was mixed with SC in

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a certain mass ratio (mass ratio of TA/SC up to 0, 0.1, 0.3, 0.5 and 1), and

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the protein concentration was 1%(w/v). The surface hydrophobicity was

159

measured after dilution to 0.1%(w/v). TA/SC solution (4mL) was mixed

160

with 40 µL ANS (8.0 mM in 0.1M PBS, pH 7.0). Fluorescence intensity

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of the samples was recorded on spectrofluorimeter (F-4600, HITACHI

162

Ltd, Japan) with a quartz cell, at λex = 390 nm, λem = 400-600 nm. So was

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evaluated by the following equation：

164

So= S2− S1

165

where S2 represents the area of the fluorescence spectrum, S1 represents

166

the area of the buffer, and relative exposed hydrophobicity(So ) was

167

expressed as S2−S1 32. The analyses were carried out in 3 repetitions.

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Foam properties. The method reported in previous study 33 was used for

169

measuring the foamability (FA) and foam stability (FS).

(3)

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Foamability (FA). 20 mL of TA/SC (mass ratio of 0, 0.1, 0.3, 0.5 and

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1) nanocomplexes solutions were foamed at 8000 rpm for 2 min with a

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homogenizer (T18, IKA) at constant temperature (25±0.2 ℃). Measuring

173

cylinder (50 mL) was carefully filled up with foam. To obtain a constant


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volume, the top of the foam was flattened with a metal spatula.

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Foamability was calculated with the following equation:

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FA(%) =

foam volume − 20 × 100 20

(4)

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Foam stability(FS). The foam volume was recorded over time. The

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foam volume at 75 min and initial time was used to evaluate the foam

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stability(FS):

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FS =

181

where V1 is the initial foam volume and V2 is the foam volume at 75 min.

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In addition, the foam size was monitored using the Dynamic Foam

183

Analyzer (DFA100FSM, Krüss GmbH, DE). According to the description

184

of measurement given by Oetjen34.

V2 × 100% V1

(5)

185

In addition, compared with the foam volume changed over 12.5 h of

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the different samples (TA/SC mass ratio of 0 and 0.3), all analyses were

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carried out in 3 repetitions.

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Dilational rheology. The surface pressure and dilational modulus for

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TA/SC nanocomplexes at air/water interface were carried out with a

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dynamic drop Tracker tensiometer (IT Concept, France). The SC

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concentration in all aqueous solutions were fixed at 1% (w/v) and the

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mass ratio of TA/SC was up to 0, 0.1, 0.3, 0.5 and 1 respectively. A

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droplet was formed (constant volume at 5 µL) by using a glass SGE

194

syringe equipped with a U-shaped metal needle and dipped into a

195

rectangular glass cuvette (25 mL) with SC or TA/SC nanocomplexes ACS Paragon Plus Environment


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196

solution. The droplet profile was continuously taken from a CCD camera，

197

then the image of the drop was digitized and analyzed. Measurements

198

were performed until it reaches a stable adsorption state (around 3h). The

199

cuvette, syringe and needle were cleaned intensively before each

200

measurement.

201

Surface pressure. The surface pressure (π) was determined by

202

analyzing the recorded droplet profile. The surface pressure is π=γo-γ,

203

where γo is the solvent interfacial tension and γ is the interfacial tension

204

of tested sample solution at adsorption time (t). The characteristic

205

adsorption time(t) and the diffusion rate constant (kdiff) were determined

206

by fitting experimental curves with a revised form of a previous Equation

207

(6)35 described by:

208

π = 2C0kT(

kdiff t 1 / 2 ) 3.14

(6)

209

Interfacial dilational properties. To obtain interfacial dilational

210

parameters, sinusoidal interfacial expansion and compression were

211

measured at appropriate frequency (f) and amplitude (dA/A) of the drop.

212

In this experiment, f is constant at 0.1 Hz (periods of 10s) and dA/A is

213

10%, which is in the range of linear viscoelasticity. The linear

214

viscoelasticity of the interfacial dilational modulus(E, Equation(9)) can

215

be defined as the ratio of the interfacial tension change(σ, Equation (7))

216

to the relative change of the interface area(A, Equation (8))36:

217

σ = σ sin(ωθ + δ ) 0


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(7)

219

A = A0 sin(ωθ )

220

(8)

221

E=

dσ dπ = = E d + iE v dA / A d ln A

222

(9) 223

where σ0 and A0 represents the stress and strain amplitudes, and δ is the

224

phase angle between stress and strain. The real part (Ed =|E| cos δ) is the

225

elastic

226

contribution of the elastic part of the viscoelastic surface. The imaginary

227

part (Ev= |E| sin δ) is a viscous component, also known as loss modulus,

228

reflecting the contribution of the viscous part for the viscoelastic surface.

229

Dilational viscoelasticity is a parameter for assessing the resistance to

230

deformation of the interfacial film. The absolute value of the dilational

231

modulus (|E|) is the total deformation resistance of the material to elastic

232

and viscous deformation.

233

Statistical analysis. A variance (ANOVA) analysis of data was made by

234

using SPSS 19.0 statistical analysis system. Significance was considered

235

at p < 0.05 throughout the study.

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RESULTS AND DISCUSSION

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Interactions of TA and SC in bulk assessed by z-Average diameter (Dz)

238

and Zeta potential measurements. Dynamic light scattering (DLS) and

239

laser Doppler velocimetry (LDV) technique have been applied in many

component

representing

storage

modulus,


reflecting

the


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37, 38

240

studies to probe protein-polyphenols interactions

241

distribution of SC and the nanocomplexes formed with different TA

242

concentrations is shown in Figure. 1A. Single SC showed a multimodal

243

distribution at pH 6.0, which was probably due to the dissolution of

244

calcium phosphate (stabilize sodium caseinate) at pH 6.0，resulting in the

245

dissociation of casein monomer from the micelle39,

246

different concentrations of TA, the intensity distributions of the mixtures

247

were also bimodal. With increasing TA concentration, the intensity of

248

minor peak (broadening from 10 to 50nm) increased. Simultaneously, the

249

intensity of major peak (broadening from 90 to 300nm) decreased. This

250

result was probably corresponding to more casein monomer linked by TA

251

at pH 6.0. According to the volume distribution data (Figure 1B), for

252

different TA concentrations, the size of TA/SC nanocomplexes retained

253

the casein monomer size, which further confirmed the above behavior. As

254

can be seen in Figure 1C, the size of TA/SC nanocomplexes decreased

255

with increase of TA concentration , probably due to the formation of

256

bridging of protein molecules after addition of TA, a stable distance

257

between the protein micelles and micelles, the average particle size is

258

significantly reduced compared with pure proteins41. The intensity size

259

distribution (including volume and number size distributions) of the main

260

peak confirmed this dependence. Figure 1C also showed the zeta

261

potential

of

TA/SC

nanocomplexes.

The


. The intensity size

40

. After adding

negative

charge

of

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262

nanocomplexes increased when the concentration of TA increased, which

263

was probably due to the coating of the TA on the surface of the

264

nanoparticles to enhance its negative charge38, 42. Simultaneously, high

265

negative charge density can be imparted due to the protonation of TA and

266

generation of oxygen centers43.

267

Effects of TA concentration on So of SC. Surface hydrophobicity(So) of

268

protein reflects the distribution of hydrophobic amino acid residues on the

269

surface of the protein. Change of So for protein will obviously affect the

270

interfacial properties of protein which play an essential role in stabilizing

271

food formulations, such as dispersions, foams, and emulsions44. Hence, it

272

is of great value to demonstrate the impact of phenolic on hydrophobic

273

amino acid residues within SC under low pH conditions at 25 °C. The

274

exposed hydrophobic groups of phenolic-treated SC were evaluated using

275

a signal-intensified fluorescence probe (ANS). As shown in Figure 2,

276

with increasing ratio of TA, So of TA/SC was dramatically lower (P

Sodium

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