Insight into the effect of microcapsule technology on the processing stability of mulberry polyphenols

Journal Pre-proof Insight into the effect of microcapsule technology on the processing stability of mulberry polyphenols Denglong Li, Mingjun Zhu, Xueming Liu, Yutao Wang, Jingrong Cheng PII:

S0023-6438(20)30132-8

DOI:

https://doi.org/10.1016/j.lwt.2020.109144

Reference:

YFSTL 109144

To appear in:

LWT - Food Science and Technology

Received Date: 4 August 2019 Revised Date:

25 December 2019

Accepted Date: 10 February 2020

Please cite this article as: Li, D., Zhu, M., Liu, X., Wang, Y., Cheng, J., Insight into the effect of microcapsule technology on the processing stability of mulberry polyphenols, LWT - Food Science and Technology (2020), doi: https://doi.org/10.1016/j.lwt.2020.109144. 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. © 2020 Published by Elsevier Ltd.

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Insight into the effect of microcapsule technology on the processing stability of

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mulberry polyphenols

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Denglong Lia,b,c, Mingjun Zhua,c,d,*, Xueming Liu b, Yutao Wanga,c, Jingrong

4

Chengb,d,* a

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College of Life and Geographic Sciences, Kashi University, Kashi 844000, China

6 b

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Sericultural & Agri-Food Research Institute, Guangdong Academy of

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Agricultural Sciences, Key Laboratory of Functional Foods, Ministry of Agriculture

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and Rural Affairs, Guangdong Key Laboratory of Agricultural Products Processing,

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Guangzhou 510610, China c

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The Key Laboratory of Biological Resources and Ecology of Pamirs Plateau in

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Xinjiang Uygur Autonomous Region, The Key Laboratory of Ecology and Biological

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Resources in Yarkand Oasis at Colleges & Universities under the Department of

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Education of Xinjiang Uygur Autonomous Region, Kashi University, Kashi 844000,

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China d

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School of Biology and Biological Engineering, Guangdong Provincial

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Engineering and Technology Research Center of Biopharmaceuticals, South China

18

University of Technology, Guangzhou Higher Education Mega Center, Panyu,

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Guangzhou 510006, People’s Republic of China

20

*

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Address: South China University of Technology, Guangzhou Higher Education Mega

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Center, Panyu, Guangzhou 510006, PR China (M. J. Zhu); 133 Yihenglu,

Corresponding author.

1

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Dongguanzhuang, Tianhe District, Guangzhou 510610, PR China (J.R. Cheng)

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Tel: +86-20-3938-0623 (M. J. Zhu); +86-20-37203765 (J.R. Cheng)

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E-mail: [email protected] (M. J. Zhu); [email protected] (J.R. Cheng)

2

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Abstract

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Polyphenols are potential food additives due to their antioxidant and pigment

28

property, although their large-scale utilization in hot processed food is not available

29

yet due to the poor processing stability. The present study investigated the effect of

30

microencapsulation strategy on the processing stability of mulberry polyphenols (MP).

31

The optimal preparation parameters for MP-β-cyclodextrin microcapsule (MPM) were

32

treated by ultrasound at 450 W, 25 ˚C for 1.5 h with a core/wall ratio of 1:6. The

33

MPM formed was verified by the UV absorption, Fourier transform infrared (FT-IR)

34

spectroscopy,

35

thermogravimetry (TG) via the shifts and intensity of the peaks. Under the optimized

36

condition, the encapsulation efficiencies of the active ingredients including total

37

polyphenols, flavonoids and anthocyanins in the MPM were above 97%; the

38

processing stability including light, thermal and storage stability of the MP were

39

remarkably improved. The above results suggest that encapsulation could be a

40

potential strategy for improving the processing stability of plant polyphenols,

41

probably leading to a more efficient application of plant polyphenols in hot processed

42

food area.

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Keywords: β-cyclodextrin, Mulberry polyphenols, Inclusion, Ultrasonography,

44

Processing stability

differential

scanning

calorimetry

1

(DSC)

and

derivative

45

1. Introduction

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Mulberry fruit, enriched in polyphenols (Wen, et al., 2019), has been verified to

47

have diversified bioactivities, such as anti-diabetes (Cao, et al., 2019; Cao, et al.,

48

2018), anti-cancer, immunoregulation (Chen, et al., 2017), etc. Nowadays, mulberry

49

polyphenols (MP), as food colorants, have been studied in a number of food systems,

50

such as fruit wine, sweets, beverage and jelly (Fazaeli, et al., 2013). For instance,

51

Tomas, et al. (2015) found that mulberry juice exhibited excellent antioxidant capacity

52

in vitro. Mulberry wine has a large amount of biological compounds, which exhibits a

53

huge development space and market potential (Wang, et al., 2015). However, the poor

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processing stability, mainly sensitive to heat and light, limits its application in foods,

55

especially hot processed foods, at a commercial scale. Meanwhile, a large amount of

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environmental factors such as temperature, pH, and oxygen are verified to be

57

destructive to the stability of MP (Xu, et al., 2019).

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Previous studies showed that MP underwent seriously degradation during hot

59

processing (Cheng, et al., 2018; Cheng, et al., 2019). Consequently, improving the

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processing stability of plant polyphenols becomes an important research direction for

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realizing its wide application in hot processed food. Microencapsulation, a popular

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technique commonly used in pharmacology and food production, has been verified to

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be effective in ameliorating the material physiochemical properties, such as solubility

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and dispensability (Mangolim, et al., 2014). Besides, several processing bottlenecks of

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bioactive compounds, such as low solubility, low stability and unpleasant taste could

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also be overcome by this technology. From this point, microencapsulation might be an 2

67

effective strategy to enhance the stability of MP, perhaps leading to a better

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processing performance.

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Cyclodextrins (CDs) possess a hydrophobic cavity, which can encapsulate

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hydrophobic components and prevent them from oxidation and thermal degradation

71

(Piletti, et al., 2019). They are made of cyclic oligosaccharides linked to glucose by

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α-(1,4)-glucosidic bonds. The hydrophobic cavity of the CDs can accommodate

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different compounds and form microcapsules by taking substances in it (Mourtzinos,

74

et al., 2008). In microcapsules, CDs is the host while the encapsulated substance is the

75

guest and they could interact with each other through van der Waals forces,

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hydrophobicity and hydrogen bond (Siripatrawan, et al., 2016). These properties make

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CDs good carriers for microcapsule, which is expected to improve the stability of MP.

78

Among all CDs, β-CD is the most widely used due to its safety, availability and

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reasonable price. Furthermore, its cavity can accommodate substances with a wide

80

molecular weight range (200 to 800 g/mol) (Szente, et al., 2004).

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Consequently, the aim of this study was to improve the processing stability of MP

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by microencapsulation. The preparation parameters of MP-β-CD microcapsule (MPM)

83

were optimized, and its formation was verified by UV scanning wavelength

84

absorption, fourier transform infrared spectroscopy (FT-IR), differential scanning

85

calorimetry (DSC) and derivative thermogravimetry (TG). Additionally, the process

86

stability, including light stability, thermal stability, and storage stability, of the formed

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complexes were also investigated.

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2. Material and methods 3

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2.1Materials and chemicals

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Mulberry juice was purchased from Guangdong Bosun Health Food Co. Ltd

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(Guangzhou, Guangdong, China) and stored at 4 ˚C. β-CD was obtained from

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Macklin Biochemical Co. Ltd (Shanghai, China) and stored at room temperature until

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use. All other chemicals and reagents were purchased from Qiyun Company

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(Guangzhou, China).

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2.2 MP preparation

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MP was prepared by X-5 resin purification according to the method of (Liu, et al.,

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2007). The obtained polyphenol elution was then freeze-dried into powder and used as

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MP.

99

2.3 The measurement of the total polyphenols, flavonoids and anthocyanins

100

The

total

polyphenol

content

was

determined

by

Folin-Ciocalteu

101

method(Alhakmani, et al., 2013) , represented by (GAE) mg/g; the total flavonoid

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content was determined by spectrophotometric colorimetry (Madaan, et al., 2011),

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represented by quercetin equivalent (QE) mg/g; the quantification of total

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anthocyanins was determined by pH differential method (Giusti & Wrolstad, 2001),

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expressed as cyanidin-3-glucoside equivalent (C3GE) mg/g.

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2.4 Preparation for MP microencapsulation

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2.4.1 Screening for the optimal method

108

To

improve

the

encapsulation

efficiency

of

polyphenols,

several

109

microencapsulation methods, including homogenization, grinding, ultra-high pressure,

110

magnetic stirring and ultrasonic wave technology, were compared (Ben Abdelkader, et 4

111

al., 2018; Dong, et al., 2017; Duan, et al., 2019; Pascual Pineda, et al., 2019; Ren, et

112

al., 2016). For homogenization treatment, 0.19±0.01g MP was added into the aqueous

113

solution of β-CD and the mixture was homogenized three times at 10000 rpm for 45 s.

114

For the other treatments, before the experiment, an aqueous solution of β-CD was

115

prepared by magnetic stirring the mixture of 0.001 mol β-CD (98.0%) and 30 mL

116

distilled water at 40 ˚C for 20 min. For grinding treatment, β-CD and MP were mixed

117

in a mortar with 10 mL distilled water and ground for 30 min. For ultra-high pressure

118

treatment, the mixture of β-CD aqueous solution and MP was packed in a plastic bag

119

and pressurized at 500 M Pa for 10 min. For magnetic stirring, the mixture was

120

prepared with a magnetic stirrer under 200 rpm for 30 min at 40 ˚C. For ultrasonic

121

treatment, the mixture of β-CD aqueous solution and MP was sealed and treated by

122

ultrasonic wave at 20±1 ˚C, 400 W for 2 h. After then, the above solutions were sealed

123

and freeze-dried into solid powder, i.e. MPM.

124

The calculation formulas of the encapsulation efficiencies are as follows: Total polyphenol encapsulation efficiency（ =

Total polyphenol content in MPM × 100% Total polyphenol content added

Total flavonoid encapsulation efficiency（ =

125 126

） %

Total flavonoid content in MPM × 100% Total flavonoids content added

Encapsulation efficiency of total anthocyanins （ =

） %

"

） %

Content of total anthocyanins in MPM × 100% Content of total anthocyanins added

2.5 Optimization of treatment parameters for MPM Ultrasonic treatment was the optimal method obtained, and the treatment 5

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parameters, including core material/wall material ratio (1:2, 1:4, 1:6, 1:8 and 1:10),

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ultrasonic time (0.5, 1.0, 1.5, 2.0 and 2.5 h), ultrasonic power (300, 350, 400, 450 and

129

500W), and ultrasonic temperature (15, 20, 25, 30 and 35 ˚C), were optimized. The

130

optimal conditions were determined based on encapsulation efficiency of the phenolic

131

compounds (total polyphenols, flavonoids and anthocyanins) by following the single

132

factor alternative method, i.e., varying one variable at a time and holding the

133

previously optimized factors constant.

134

2.6 Characterization of microcapsules

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2.6.1 UV scanning wavelength absorption

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Sample of 0.01 g was dissolved in 10 mL ultrapure water. After that, the

137

ultraviolet absorption spectrums of MP, β-CD and MPM were obtained using a

138

UV-Vis spectrophotometer (UV-1800, SHIMADZU, Japan), respectively, and their

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maximum absorption wavelength were recorded.

140

2.6.2 FT-IR spectra

141

The FT-IR spectra of the samples were obtained using an infrared fourier

142

transform spectrometer (model Vertex 70v, Bruker, Germany). The spectral range was

143

400-4000 cm-1 with 128 scans and a resolution of 2 cm-1. The samples were diluted in

144

potassium bromide (KBr) powder and the pellets formed were used for analysis.

145

2.6.3 DSC analysis

146

The DSC thermograms of the samples were measured with a DSC 200 F3

147

(NETZSCH, German). The scanning temperature range is 30~300 ˚C with a heat rate

148

of 10 ˚C /min under nitrogen atmosphere. 6

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2.6.4 TG analysis

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A thermogravimetric analyzer STA449 F3 Jupiter (NETZSCH, German) was used

151

to determine the thermal properties and behavior of the MP, β-CD, the mixture of MP

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and β-CD, and MPM. N2 was used as the carrier with a flow rate of 40 mL/min. The

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heating temperature was ranged from 30  to 900  ˚C with heating rate of 10 °C/min.

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2.7 Stability of MPM

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2.7.1 Light stability analysis

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The photochemical stability of MP and MPM was assessed with fluorescent

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lighting at room temperature. To be more specific, 5 g MP and MPM was exposed to

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fluorescent lighting, respectively, for 48 h in enclosed glass petri dishes (30×30 mm).

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After 0, 12, 24, 36 and 48 h, samples were collected, and the MP retention rates were

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recorded.

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2.7.2 Thermal stability

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The thermal stability of MPM was assessed according to the method described by

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Paramera, et al. (2011) with a minor modification. Isothermal heating was conducted

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under oxidative conditions. During the process, 5 g MP and MPM were heated by

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water bath at 25, 35, 45, 60 and 100 ˚C for 60 min, respectively. After that, the

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retention rate was determined.

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2.7.3 Storage stability

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Storage stability of MPM was evaluated by analyzing the degradation of the

169

microcapsules at room temperature (25±2 ˚C). To be specific, the present study

170

compared the degradation of polyphenols in MPM during 28 days of storage under 7

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two packaging methods (no packaging and vacuum packaging). The samples were

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collected every 7 days and retention rates were recorded by spectrophotometric

173

analysis.

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2.8 Statistic analysis

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Each experiment was done in triplicate with data reported as mean and standard

176

deviation. The analyses were performed using the SPSS version 17.0 software for

177

windows (SPSS Inc., Chicago, Illinois). ANOVA and Duncan’s multiple range tests

178

were conducted to determined significant differences, and a value of P<0.05 was

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considered statistically significant.

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3. Results and discussion

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3.1 Screening of the optimal method for MPM preparation

182

Encapsulation efficiency directly reflects the effectiveness of wall material

183

(Ahmad, et al., 2018). In this study, the total polyphenols, flavonoids and

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anthocyanins in MP were 406.00±1.36 mg GAE /g, 94.47±1.08 mg QE/g and

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73.59±1.25 mg C3GE/g, respectively. The treatments showed significant effect on the

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encapsulation efficiencies of phenolic compounds (Fig. 1). Encapsulation efficiency

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of total polyphenols (EETPE%) and flavonoids (EETFE%) exhibited a similar

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encapsulation efficiency ranged from 70.0% to 89.0%, while encapsulation efficiency

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of anthocyanin (EETAE%) were 77.0%~90.0%. Notably, the highest encapsulation

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efficiency (EETPE% 89.8%; EETFE% 89.9%; EETAE% 91.0%) was obtained in sample

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treated with ultrasonic wave. This phenomenon indicates that β-CD is an effective

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film-forming agent for MP embedding (Ahmad, et al., 2017; Akhavan Mahdavi, et al., 8

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2016). Similarly, Mangolim, et al. (2014) also claimed that β-CD was a potential wall

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material for curcumin microcapsulation and they reported an encapsulation efficiency

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of 74% in curcumin-β-CD complex prepared by co-precipitation. Due to the highest

196

encapsulation efficiency, ultrasonic treatment was chosen for MPM preparation in the

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further study.

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3.2 Optimization of preparation parameters for MPM

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3.2.1 Core/wall ratio

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As shown in Fig. 2 (A), all encapsulation efficiencies, including EETPE%, EETAE%

201

and EETFE%, exceeded 60.0% when core/wall ratio ranged from 1:2 to 1:10. EETPE%

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increased progressively (P<0.05) as the core/wall ratio increased from 1:2 to 1:6, and

203

the highest encapsulation efficiency (EETPE%: 89.8%; EETFE%: 89.8%; EETAE%:

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90.9%) was obtained with a core/wall ratio of 1:6. However, the increment terminated

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when the core/wall ratio further increased. This is perhaps ascribed to the changes in

206

physicochemical property of the solution. As Xu, et al. (2019) claimed that excessive

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β-CD increased the solution viscosity, thus resulting in a poor dispersion of β-CD. In

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addition, excessive core material content may reduce the intensity of the wall material,

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thus impacting the encapsulation efficiency (Xu, et al., 2019). Based on the

210

encapsulation efficiency result, 1:6 was determined as the optimal core/wall ratio for

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MPM preparation.

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3.2.2 Ultrasonic time

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The effect of ultrasonic time on encapsulation efficiency of polyphenols is shown

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in Fig. 2B. With the extension of ultrasonic time, the polyphenol encapsulation 9

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efficiency increased steadily and reached the highest value with an ultrasonic time of

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1.5 h (EETPE% 94.0%, EETFE% 92.3% and EETAE% 93.4%). However, further prolong

217

the ultrasonic time, the increasing trend terminated. This was perhaps due to the

218

dissociation of the resultant microcapsules (Sun, et al., 2018).

219

3.2.3 Ultrasonic power

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Parallel to the variations of core/wall ratio and ultrasonic time, encapsulation

221

efficiency of the polyphenols also presented a trend of first rising and then falling. As

222

shown in Fig. 2 (C), the highest encapsulation efficiencies (EETPE% 95.5%, EETFE%

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94.7% and EETAE% 96.2%) were obtained with an ultrasonic power of 450 W. As

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claimed by Silva, et al. (2015), high-intensity sound waves could intrigue ultrasonic

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"cavitation effect", which generated energy intensification and bursts instead of

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agitating the mixture. Consequently, further increasing the ultrasonic power could

227

re-intensify MP and reduce its encapsulation efficiencies. Therefore, the 450 W was

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selected for MPM preparation in further study.

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3.2.4 Ultrasonic temperature

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Ultrasonic temperature also exhibited pronounced effect on the encapsulation

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efficiencies of phenolic compounds. As shown in Fig. 2(D), the encapsulation

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efficiencies of MPM increased smoothly with the increasing of temperature and the

233

highest encapsulation efficiency (EETPE% 97.2%, EETFE% 97.2% and EETAE% 97.2%)

234

was obtained with an ultrasonic temperature of 25 ˚C. This is because the increasing

235

of temperature enhances the solubility of MPM in the given solution. Higher

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temperature accelerates the movement of molecules and jeopardizes the crystal lattice 10

237

stability, leading to effective collisions between β-CD and the MP, thus improving the

238

encapsulation effectiveness of the phenolic compounds (M. Liu, et al., 2015).

239

However, if the temperature exceeds a certain limit, the equilibrium could be

240

destroyed, and the non-covalent bond between β-CD and MP cracked, resulting in the

241

breakdown of the synthesized microcapsules. Notably, there was no statistical

242

difference in the encapsulation efficiencies of MPM prepared with an ultrasonic

243

temperature range of 20~30 ˚C. Since 25 ˚C is close to room temperature and no

244

heating or cooling is required, this temperature was selected for MPM preparation.

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3.3 Characterization of MPM

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3.3.1 UV analysis

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The maximum UV absorption wavelengths of β-CD, MP and MPM were

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presented in Fig. 3(A). Apparently, there was no statistically significant difference in

249

the maximum absorption wavelength (519.5 nm) between samples of MP and MPM.

250

No significant absorption was detected in β-CD within the ultraviolet wavelength

251

scanning range. However, it has been noted that, compared with MP, the absorption of

252

MPM at 519.5 nm was remarkably reduced. The recurrence of the maximum

253

absorption wavelength of core material in microcapsule and attenuation of absorption

254

values also reported by other researchers (Maisuthisakul, et al., 2012; Rodrigues, et al.,

255

2011), which suggested that the core material has successfully entered the cavity of

256

β-CD.

257

3.3.2 FT-IR

258

The spectrums of the MP, β-CD, the mixture of MP and β-CD and MPM are 11

259

shown in Fig. 3 (B). β-CD showed characteristic bands at 3361 cm-1 (-OH stretching),

260

2923 cm-1 and 577 cm-1 (C-H stretching), 1158 cm-1 (C-C stretching), 1081 cm-1and

261

1028 cm-1 (C-O stretching). Meanwhile, MP showed characteristic bands at 3340 cm-1

262

(O-H stretching), 2961 and 2925 cm-1 (C-H stretching), 1605 cm-1 (C=C stretching of

263

aromatic ring), 1259 cm-1 (C-O stretching) and 1020 cm-1 (C-H deformation of

264

aromatic ring)(Peralta, et al., 2019). When MP and β-CD mixed, the above

265

characteristic peaks appeared again. However, different peak profiles occurred in the

266

FT-IR spectrum of the MPM though most characteristic peaks of MP and β-CD

267

reappeared. In particular, all characteristic peaks of the β-CD reoccurred while the

268

characteristic peaks of MP at 2961 cm-1, 2925 cm-1 and 1259 cm-1 disappeared in the

269

FT-IR spectrum of the MPM. The above results indicate that the benzene ring of MP

270

have successfully entered the cavity of β-CD and a new compound have been formed

271

(Aigner, et al., 2012).

272

3.3.3 DSC analysis

273

The inclusion effect between host and guest molecules can trigger the absence or

274

movement of endothermic peaks, reflected by changes in crystal melting, boiling or

275

sublimation points (Horvath, et al., 2008). As shown in Fig 3 (C), β-CD had a wide

276

heat absorption peak between 50-190 °C, of which the maximum absorption occurred

277

at 141.93 ˚C. The endothermic peak of MP was between 50-119.40 ˚C, of which the

278

maximum absorption appeared at 94.12 ˚C. When they mixed, its absorbance shifted

279

to 131.97 ˚C, which implied the interaction between β-CD and MP. Notably, their

280

absorbance peak was quite different from the MPM, of which a new endothermic 12

281

peak appeared at 214.57 ˚C. Similar to the result of FI-RP, the appearance of the new

282

peak indicates that MP molecule has entered the β-CD cavity successfully and the

283

inclusion complex has formed. This result was identical to that of Sousdaleff, et al.

284

(2013) who claimed that the movement or disappearance of the melting point, boiling

285

point and sublimation point predicted the formation of a new complex.

286

3.3.4 TG analysis

287

The thermal stability of MP, β-CD, mixture and MPM were studied by TG. As can

288

be seen from Fig. 3(D), pyrolysis process of β-CD consists of two stages, of which

289

peaks appeared at 112 ˚C and 400 ˚C, respectively. To be specifically, the weight loss

290

rate of 13.0% occurred in the first stage, which was the process of losing crystal water;

291

another weight loss (77.6%) appeared in the second storage, which was due to the

292

thermal decomposition of β-CD. Unlike β-CD, decomposition of MP occurred with

293

the increase of temperature, and a weight loss of 58.8% was detected during

294

138-600 °C. The thermogravimetric behavior of the physical mixture was the

295

superposition of MP and β-CD. It is noteworthy that the thermogravimetric curve of

296

MPM was rather different from the other three samples. More precisely, a small

297

weight loss (11.1%) occurred at the first stage (60-180 ˚C), while a larger loss (65.7%)

298

appeared at the second stage (260-460 ˚C). This different behavior indicates the

299

formation of a new complex.

300

3.4 Stability analysis of MPM

301

3.4.1 Light stability

302

Light has a great influence on the stability of MP. After 48 hours illumination, 13

303

phenolic compounds underwent significant degradation (Fig. 4). In MP, the contents

304

of total polyphenols, flavonoids and anthocyanins lost 38.6%, 14.9% and 39.0%,

305

respectively. Specifically, the retentions of total polyphenols, flavonoids, and

306

anthocyanins in MPM were 59.7%, 82.8% and 58.5%, which were 1.34, 1.05 and

307

1.36-fold of that in the MP, respectively. These results indicate that the light stability

308

of MP is enhanced by microencapsulation with β-CD. Similar to the present study,

309

Munhuweyi, et al. (2018) and Woranuch, et al. (2013) also suggested that the

310

polyphenol microcapsules prepared with CDs showed good antioxidant stability and

311

thermal stability. This is because CDs are cyclic oligosaccharides with a stepped

312

hollow vertebral body and are composed of glucose monomers connected by α-(1.4)

313

bonds. CDs can form noncovalent object-guest complexes with several molecules

314

including essential oils, fragrances/flavors, and antioxidants (Kayaci, et al., 2014).

315

Similarly, Feng, et al. (2019) also reported an improved decomposition temperature of

316

the 1, 2-O, O-diacetyllycorine after microencapsulation processing with α-CD.

317

3.4.2 Thermal stability

318

Fig. 5 depicts the thermal stability results of MP and MPM between 25-100 ˚C.

319

The loss of phenolic compounds, due to polymerization and oxidative degradation,

320

occur during thermal processing (You, et al., 2018). Under lower temperature (20 ˚C)

321

polyphenols compounds exhibited less loss (12.0%) in both MP and MPM. Increased

322

temperature significantly reduced the retention of polyphenols and anthocyanins.

323

Especially, when temperature exceeded 45 °C, the rising of temperature increased the

324

flavonoid retention. This is probably because high temperature destroys the structure 14

325

of flavonoids, exposing more phenolic hydroxyl structures, which thus lead to the

326

increase of absorption value (Lu, et al., 2018). In general, the stability of phenolic

327

compounds in MPM was significantly stronger than that in MP. This is mainly

328

ascribed to the physical protection barrier of β-CD (Piletti, et al., 2019). Moreover, the

329

complexation effect between the β-CD and MP may also be responsible for the

330

increased stability (C. S. Mangolim, et al., 2014).

331

3.4.3 Storage stability

332

The storage stability of MP and MPM was assessed for 28 days under vacuum

333

packaged and unpackaged conditions (Fig.6). All tested phenolic compounds

334

underwent degradation during storage no matter whether vacuum-package was used.

335

More than 44.0% total polyphenols, 20.0% flavonoids and 40.0% anthocyanins lost

336

during the storage period. Similar to the results obtained in light and thermal stability

337

tests, the phenolic compounds in MPM were more stable than those in MP during the

338

whole storage stage. A typical example could be found in samples without vacuum

339

package, of which the total polyphenols, flavonoids and anthocyanins retention ratios

340

were 65.9%, 77.7% and 59.1% in MP while those in MPM increased to 70.4%, 79.2%

341

and 68.0% within 28 days storage, respectively. Our findings coincide with the result

342

of Paramera, et al. (2011), who also claimed that the storage stability of curcumin

343

could be enhanced after microencapsulated by β-CD and modified starch. Moreover,

344

Ho, et al. (2017) found that microencapsulation with β-CD could protect catechin

345

against temperature, light and oxygen. At the end of the storage period (day 28), 79.2%

346

polyphenols, 88.0% flavonoids and 78.2% anthocyanins were obtained in MPM 15

347

vacuum-packaged , while only 70.4% polyphenols, 79.2% flavonoids and 68.0%

348

anthocyanins were detected in MPM without package. This is mostly due to the

349

oxidative decomposition of polyphenols (J. R. Cheng, et al., 2016). From the above

350

analysis, blocking oxygen is still an effective method to prevent the phenolic

351

compounds from degradation, which should be taken account for the future study.

352

4. Conclusion

353

The microcapsules of MP were successfully prepared by ultrasonic method with

354

the wall material of β-CD. The optimal preparation parameters for MPM were ultra

355

sound at 450 W, 25 ˚C for 1.5 h with a core/wall ratio of 1:6. Under the optimized

356

condition, the encapsulation efficiencies of the active ingredients, including total

357

polyphenols, flavonoids and anthocyanins, were above 97.0%, and the processing

358

stability, including light stability, thermal stability, and storage stability, of the MP

359

were remarkably improved. This study developed a novel method to improve the

360

stability of mulberry polyphenols, which is expected to make better use of plant

361

polyphenols in the field of food processing.

362

Acknowledgement

363

The authors gratefully acknowledge the financial support of Province Natural

364

Science Fund of Guangdong [grant numbers 2018A030313202; 2018A030313796];

365

Guangzhou Science and Technology Program key projects [grant numbers

366

201807010080, 201806040007, 201704020054]; Guangdong Science and Technology

367

project [grant numbers 2017A040405036; 2018LM2154]; Guangdong Yangfan

368

Program [grant number 2016YT03H079]; R&D Projects in Key Areas of Guangdong 16

369

Province [2019B020212003].

370

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Fig.1. Effects of preparation methods on the encapsulation efficiency of the phenolic compounds.

: Total anthocyanins,

: Total polyphenols,

: Total

flavonoids. Values with different letters (a-c) in the same index are significantly different (P< 0.05).

Fig.2. Preparation parameter optimization of MP-β-CD microcapsule (MPM). (A) Mass ratio of mulberry polyphenols (MP) to β-cyclodextrin (β-CD); (B) Ultrasonic time; (C) Ultrasonic power; (D) Ultrasonic temperature. content,

: Total polyphenols content,

: Total anthocyanins

: Total flavonoids content. a-d:

Different letters above bars indicate significant differences between various treatments of the same index (P<0.05).

Fig.3. Validation analysis of microcapsules, UV absorption curves (A), FT-IR spectra (B), DSC thermograms of samples (C) and TG curves (D).

Fig.4. Comparison of the light stability between MP and MPM. anthocyanins in MPM, MPM,

:Total anthocyanins in MP,

:Total polyphenols in MP,

:Total

: Total polyphenols in

:Total flavonoids in MPM,

:Total

flavonoids in MP. a-e: Values with different letters within the same treatment are significantly different (P< 0.05).

Fig.5.Comparison of the thermal stability between MP and MPM. anthocyanins in MPM, MPM,

: Total anthocyanins in MP,

: Total polyphenols in MP,

:Total

: Total polyphenols in

: Total flavonoids in MPM,

: Total

flavonoids in MP. a-e: Values with different letters within the same treatment are significantly different (P< 0.05).

Fig.6. Comparison of the storage stability between MP and MPM. (A) Total polyphenols; (B) Total flavonoids; (C) Total anthocyanins. natural conditions, natural conditions,

:MP stored in vacuum conditions, :MPM stored

: MP stored in :MPM stored in

in vacuum. a-e: Values with different letters

within the same treatment are significantly different (P< 0.05).

Mulberry polyphenol microcapsule was constructed with β- cyclodextrin. The processing parameters of mulberry polyphenol microcapsule were optimized The light stability of mulberry polyphenol was improved by microcapsule technique. The thermal stability of mulberry polyphenol was improved by microcapsule technique. The storage stability of mulberry polyphenol was improved by microcapsule technique.

Dear Editor, This manuscript is an original research paper of authors. Neither the entire paper nor any part of its content has been submitted to LWT – Food Science and Technology earlier or any other journal. The submission to LWT – Food Science and Technology is also approved by all authors. And if accepted, it will not be published elsewhere including electronically in the same form, in English or in anyother language, without the written consent of the copyright holder. Yours Sincerely, Jingrong Cheng

Conflict of interests

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.