Accepted Manuscript

High methoxyl pectin from dragon fruit (Hylocereus polyrhizus) peel

Kharidah Muhammad, Nur Izalin Mohd. Zahari, Sri Puvanesvari Gannasin, Noranizan
Mohd. Adzahan, Jamilah Bakar

PII: S0268-005X(14)00094-0
DOI: 10.1016/j.foodhyd.2014.03.021
Reference: FOOHYD 2548

To appear in: Food Hydrocolloids

Received Date: 31 January 2013

Accepted Date: 19 March 2014

Please cite this article as: Muhammad, K., Izalin Mohd. Zahari, N., Gannasin, S.P., Adzahan, N.M.,
Bakar, J., High methoxyl pectin from dragon fruit (Hylocereus polyrhizus) peel, Food Hydrocolloids
(2014), doi: 10.1016/j.foodhyd.2014.03.021.

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Whole peel/Epicarp • Pectin yield: 26.38%

• Degree of esterification:
Inner layer of peel
63.74%

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Peel with inner • Molecular weight:
layer removed 0.88 x 105 Da

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Dragon fruit

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1 High methoxyl pectin from dragon fruit (Hylocereus polyrhizus) peel

3 Kharidah Muhammad*, Nur Izalin Mohd. Zahari, Sri Puvanesvari Gannasin, Noranizan Mohd.

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4 Adzahan, Jamilah Bakar

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6 Faculty of Food Science and Technology, Universiti Putra Malaysia, 43400 UPM, Serdang,
7 Selangor Darul Ehsan, Malaysia

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16 *Corresponding author. Tel.: +603-89468394; Fax: +603-89423552.

17 E-mail address: kharidah@putra.upm.edu.my (K. Muhammad).

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26 ABSTRACT

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28 Pectin from different fractions of dragon fruit (Hylocereus polyrhizus) peel was extracted using

29 1% citric acid and the physico-chemical characteristics of the pectin were studied. The highest

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30 pectin yield (26.38% on dry weight basis) was obtained from fresh inner layer of the peel when

31 extraction was carried out at temperature: 73 oC, time: 67 min, pH: 2.03, and sample to citric acid

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32 ratio: 1:4 (w/v). The pectin also demonstrated the highest degree of esterification (63.74%) when

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33 compared with pectin from other fractions of the dragon fruit peel investigated in this study. The

34 calculated degree of esterification confirmed that the extracted pectin is a high methoxyl pectin.

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35 The molecular weight of the pectin determined using size exclusion chromatography was 0.88 x
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36 105 Da. Monosaccharide composition determined using high performance liquid chromatography

37 revealed that the pectin was predominantly constituted of galacturonic acid (39.11%), followed
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38 by moderate concentrations of mannose, rhamnose, galactose, glucose and minor amounts of

39 xylose and arabinose. The pectin exhibited Newtonian behaviour at concentrations of 0.5% and
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40 1.0%, and pseudoplastic behaviour at concentrations of 2.0% and 3.0%. Although the viscosity
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41 of the dragon fruit peel pectin was lower than that of commercial apple and citrus pectins, it can
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42 be used as a functional and health ingredient in low viscous foods and beverages.

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44 Keywords: Extraction; physico-chemical; characterisation; dragon fruit peel; high methoxyl

45 pectin
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51 1. Introduction

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53 Dragon fruit or pitaya, a member of Cactaceae is native to tropical regions of Mexico,

54 and Central and South America (Mizrahi, Nerd, & Nobel, 1997). It is also widely grown in Asian

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55 countries such as Malaysia, Vietnam, Thailand, the Philippines and Taiwan (Mizrahi, Nerd, &

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56 Nobel, 1997; Nerd et al., 2002; Wu et al., 2006). Different varieties of dragon fruits have been

57 developed as fruit crops such as Hylocereus undatus (red epicarp, white pulp), Hylocereus

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58 megalanthus (yellow epicarp, white pulp), Hylocereus polyrhizus (red epicarp, red-purple pulp)

59 and Hylocereus costaricensis (red epicarp, red pulp) (Ariffin et al., 2009; Esquivel et al., 2007;

60 Nerd et al., 2002).

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61 Recently, H. polyrhizus has received much attention as a source of natural red-purple
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62 colour that has great potential in colouring a broad array of foods (Esquivel et al., 2007; Syed

63 Muhammad et al., 2011; Wybraniec & Mizrahi, 2002). In addition, H. polyrhizus is also very
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64 well known for its good antioxidant activity that may exert various health benefits (Tenore et al.,
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65 2002; Wu et al., 2006). Besides the added-value natural colourant, the pulp of H. polyrhizus is

66 also processed into nutritional juice in order to increase the commercialisation value of the fruit.
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67 The utilisation of the fruit pulp for the production of natural colouring agent (Syed
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68 Muhammad et al., 2011) and juice (Herbach et al., 2007) has indirectly resulted in the
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69 accumulation of the fruit peels as waste materials. Therefore, this study is dedicated to extract

70 and characterise pectin from H. polyrhizus peel aiming at adding value to the disposed peel.

71 Pectin is a high-value functional ingredient widely used as gelling agent in jams, and as

72 stabiliser in acidified dairy drinks (May, 2000). Pectin is a family of galacturonic acid-rich

73 polysaccharides including homogalacturonan, rhamnogalacturonan I, rhamnogalacturonan II, and

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74 xylogalacturonan (Mohnen, 2008). Depending on the degree of esterification (DE), pectin is

75 divided into two major groups: pectins with DE > 50% are known as high methoxyl pectins

76 whereas low methoxyl pectins have a DE < 50% (Morris et al., 2000). Apple pomace and citrus

77 peels are the major sources of commercially available pectins (Thakur et al., 1997). The

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78 composition of pectins actually varies depending on the plant source and extraction conditions

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79 (Rolin, 1993). Thus, the physico-chemical characteristics of pectin extracted from H. polyrhizus

80 peel were studied.

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82 2. Materials and methods

83 2.1. Materials
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84 Dragon fruits with red epicarp and red-purple pulp (H. polyrhizus) were obtained from a
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85 fruit farm in Selangor, Malaysia. Monosaccharide, dextran standards, citrus and apple pectins

86 were purchased from Sigma-Aldrich (USA). Citrus pectin with galacturonic acid content ≥ 74%
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87 and apple pectin with degree of esterification 70-75% as specified by the supplier were used.
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88 HPLC grade chemicals used for monosaccharide profiling were purchased from Merck

89 (Germany). Other chemicals used in this experiment were of analytical grade.

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90 2.2. Sample preparation

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91 The whole peel (epicarp) of each dragon fruit was separated from the pulp immediately
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92 after the arrival of the fruits in the laboratory conditioned at 25 oC. Inner layer of the peel was

93 also obtained after removal of the outer layer of the peel. The whole peels and the inner layer of

94 the peels were cut into small pieces and dried in an air-circulated oven (Venticell, Medcenter

95 Einrichtungen GmbH, Germany) at 55 oC until constant weights were obtained. The dried peels

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96 were then ground using a pulverisette 14 variable speed rotor mill (Fritsch GmbH, Oberstein,

97 Germany). The dried powder obtained using the whole peels and the inner layer of peels were

98 identified as WPD and IPD, respectively. Fresh whole peels and inner layer of peels were also

99 homogenised with increasing speed repeatedly (4 x 1 min) using a Waring laboratory blender

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100 (MS 153-5, Torrington, USA) and were identified as WPF and IPF, respectively. Either one of

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101 the two options for sample preparation (dry/fresh form) can be considered by the pectin

102 manufacturers depending on their sample storage facility, stability and cost.

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103 2.3. Pectin extraction

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104 2.3.1. Pectin extraction from dried dragon fruit peels
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105 Dragon fruit peel powder (WPD or IPD) was mixed with 1% citric acid (1:50 w/v) at pH

2.03 in a 250 ml Schott bottle using a shaking water bath. The extraction was carried out at 73 oC
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107 for 67 min. The resulting extract was cooled to room temperature (25 oC) and filtered through a
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108 Whatman No. 113 filter paper using a Buchner funnel which was connected to a vacuum pump
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109 (Mollea et al., 2008). The filtrate was then concentrated to half of its initial volume with a rotary

110 evaporators set at 52 ± 1 oC (Favarash & Ashtiani, 2007). For precipitation of the pectin, two
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111 volumes of 95% undenatured ethanol were added to one volume of the filtrate with gentle

112 stirring to break up the forming gelatinous lumps (Masmoudi et al., 2008). The mixture was kept
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113 for 1 hr at 4 oC to allow pectin floatation and to reach equilibrium colloid-liquid state (Favarash
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114 & Ashtiani, 2007). The floating pectin was then separated by filtration through a Whatman

115 No.113 filter paper and washed three times with 50%, 75% and 100% ethanol, sequentially. The

116 extracted pectin was then dried in an air-circulated oven at 50 oC for 15 hr and weighed. The

117 dried pectin was ground using a pulverisette 14 variable speed rotor mill (Fritsch GmbH,

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118 Oberstein, Germany). The pectin yield when dried dragon fruit peel was used (Yd) was

119 calculated using equation 1:

120 Yd (%) = 100 [weight of dried pectin (g)/weight of dragon fruit peel powder (g)] (1)

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121 2.3.2. Pectin extraction from fresh dragon fruit peels

122 Fresh dragon fruit peel (WPF or IPF) was mixed with 1% citric acid (pH 2.03) using 1:4

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123 (w:v) sample to citric acid ratio in a 250 ml Schott bottle using a shaking water bath. The

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124 extraction was carried out at 73 oC for 67 min. The extraction temperature, time and pH were

125 similar as used in subsection 2.3.1., only the sample to extracting solution ratio was different

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126 where 1:4 (w/v) was used instead of 1:50 (w/v) since the moisture content of fresh dragon fruit
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127 peels is > 90%. The extraction method from here on, however, is as described by Yuliarti et al.

128 (2008) whereby the resulting extract was centrifuged at 3,300 x g for 20 min and the supernatant
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129 was decanted. Citric acid (1%) at 73 ºC was added to the pellet in the ratio of 1:1 (w:v) to extract
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130 any remaining pectin, and centrifuged again as before. Undenatured ethanol (95%) was added to
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131 the combined supernatants, the mixture was stirred and kept at 4 ºC for 4 h to facilitate pectin

132 precipitation. The mixture was centrifuged at 3,300 x g for 10 min and the supernatant was
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133 discarded. The pellet was washed with 95% undenatured ethanol (1:1; w:v) and centrifuged

134 again. This pellet washing step was further repeated. The final pellet was dried, weighed and
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135 ground as described in subsection 2.3.1. The pectin yield on fresh (Yf) and dry (Y) weight basis
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136 when fresh dragon fruit peel was used were calculated using equations (2) and (3), respectively:

Yf (%) = 100 [weight of dried pectin (g)/weight of fresh dragon fruit peel (g)] (2)

Y (%) = 100Yf /(100-moisture content of fresh dragon fruit peel) (3)

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137 2.4. Functional groups determination using Fourier transform infrared (FT-IR) spectroscopy

138 Samples were desiccated overnight in a glass dessicator containing anhydrous silica gel

139 prior to FT-IR analysis. FT-IR spectra were recorded using a universal ATR accessory on a

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140 Perkin Elmer Spectrum 100 FT-IR spectrophotometer at the absorbance mode from 4,000 to 400

141 cm-1 (mid infrared region). The measuring resolution was 4 cm-1 and 128 scans were collected to

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142 get a high signal-to-noise ratio.

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143 2.5. Determination of degree of esterification

144 The degree of esterification (DE) was determined using the FT-IR method. DE (%) is

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defined as (number of esterified carboxylic groups/number of total carboxylic groups) x 100.
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146 The ratio of the area of the band at 1,745 cm-1 (corresponding to the number of esterified

carboxylic groups) over the sum of the areas of the bands at 1,745 cm-1 and 1,630 cm-1
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148 (corresponding to the number of total carboxylic groups) should be proportional to the DE, i.e.
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149 DE = A1745/(A1745+A1630) (Chatjigakis et al., 1998; Manrique & Lajolo, 2002). OMNIC
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150 software version 8.0 (Thermo Nicolet Corp., USA) was used to measure the areas of interest.

2.6. Proximate analyses

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152 Moisture content was determined according to AOAC Method 934.01 (AOAC, 2005).
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153 Ash was analysed gravimetrically by incineration of sample in a muffle furnace for 2 h at 900°C.
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154 The protein content estimated as % nitrogen x 6.25 (conversion factor) was determined with the

155 Kjeldahl method on a KjeltecTM 8400 Analyser (FOSS-Tecator AB, Hoganas, Sweden) which is

156 consistent with AOAC Method 978.04 (AOAC, 2005). Dietary fibre was determined using

157 Megazyme Total Dietary Fibre Assay kit which is in accordance with AOAC Method 991.43

158 (AOAC, 2005) using a Fibretec System E1023 (FOSS-Tecator AB, Hoganas, Sweden).

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159 2.7. Surface morphology analysis

160 Sample was mounted onto a scanning electron microscope (SEM) specimen stub with a

161 carbon double-sided tape prior to coating. Sample was coated with gold using a Bal-Tec SCD

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162 005 sputter coater (Bal-Tec AG, Liechtenstein) and subsequently photographed using a Leo 1455

163 variable pressure SEM (Germany) at x25 and x100 magnifications.

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164 2.8. Homogeneity and molecular weight determination using size exclusion chromatography

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165 The molecular weight determination by laser light scattering was measured using the

166 method described by Corredig et al. (2000). Dragon fruit peel, apple and citrus pectins were

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168 Heleos multiangle laser light-scattering (MALLS) detector (Wyatt Technology, USA) and a
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169 Optilab reX differential refractometer (RI) which operated at 633 nm at a constant temperature,

170 25 oC. The pectin solutions (1.5 mg/mL) were solubilised under magnetic stirring, then filtered
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171 through a 0.45 µm membrane filter (Milipore Co., Milford, USA) and manually injected through
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172 a 100 μL loop. One PL aquagel-OH MIXED-H 8 μm column (Agilent Technologies, USA) was

173 used. Elution was carried out at a flow rate of 0.6 ml/min with 0.1 M sodium nitrite (NaNO2)
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174 solution containing 0.5 g/L sodium azide (NaN3) as a bactericide at 25 oC. Monodisperse dextran

175 standards with molecular weights 25, 60 and 500 kDa were used to check the performance of the
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176 HPSEC-MALLS system. Molecular weight parameters were calculated using the Astra software.
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177 2.9. Determination of monosaccharide composition

178 Monosaccharide composition of dragon fruit peel pectin was determined using the

179 method described by Dai et al. (2010) using high performance liquid chromatography (HPLC).

180 The pectin (3mg/mL) was dissolved in distilled purified water. The pectin solution (100 uL) was

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181 hydrolysed to monomers with 100 uL of 4 M trifluoroacetic acid for 2 hr in an oven at 100 ºC

182 prior to derivatisation with 1-phenyl-3-methyl-5-pyrazolone (PMP). Ten monosaccharides

183 (mannose, ribose, rhamnose, glucuronic acid, galacturonic acid, glucose, galactose, xylose,

184 arabinose, fucose) were separated using ZORBAX Eclipse Plus-C18 HPLC column (250 mm

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185 length, 4.6 mm internal diameter and 5 µm particle size, Agilent Technologies, USA). A Waters

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186 e2695 HPLC equipped with a Waters 2489 UV/Vis detector was used. The HPLC conditions and

187 mobile phase composition were as described by Dai et al. (2010). An Empower2 software

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188 (Waters, USA) was used to collect the data. The monosaccharide concentration in dragon fruit

189 peel pectin was calculated by comparing integration values of their peak areas to a calibrated

190 standard curve.

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191 2.10. Flow analysis
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192 Dragon fruit peel, apple and citrus pectins dispersions with different concentrations
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193 (0.5%, 1%, 2% and 3%, w/v) were prepared with distilled deionised water. Flow behaviour of
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194 the dispersions was determined using a rheometer (Rheostress 600, Karlsruhe, Germany).

195 Measurements were performed with a 35 mm in diameter serrated parallel plate geometry
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196 (PP35Ti). The shear rate was increased from 0 to 400 s-1 using the software control (Monsoor,

197 2005). The temperature was maintained at 25±0.1 °C throughout the measurement.
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198 2.11. Statistical analysis

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199 Statistical analysis was performed using Minitab version 14 software (Minitab Inc., State

200 College, PA, USA). Analysis of variance (ANOVA) was applied to determine significant

201 difference at p<0.05.

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203 3. Results and discussion

204 3.1. Pectin yield

205 The optimum extraction conditions (temperature: 73 ºC, time: 67 min, pH 2.03) were

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206 obtained from a preliminary optimisation study using response surface methodology where

207 twenty treatments were assigned based on the central composite design (Table 1). The extraction

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208 optimisation study was performed using the fresh inner layer of peel (IPF). Pectin extraction was

209 also carried out using the optimum extraction conditions as mentioned above for other peel

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210 fractions (WPD, IPD and WPF). As shown in Fig. 1, the pectin yield increased as the pH was

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211 decreased from 4 to 2. Commercially, pectin is extracted using hot dilute mineral acid at pH~2
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212 (May, 1990). At hold temperature of 67.5 ºC, lower pH value and longer extraction time

213 significantly increased the pectin yield. Pectin yield was not significantly (p > 0.05) affected by
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214 drying of the dragon fruit whole peels as shown by the results obtained with WPD and WPF in

215 Table 2. The pectin yield (dry weight basis) when whole peels were used was almost 15% which
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216 was consistent with that of previous findings. Ismail et al. (2012), Tang et al. (2011) and Woo et
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217 al. (2010) reported 14.96% (extracted with 0.03M hydrochloric acid), 10.40-16.76% (extracted

218 with 40% citric acid), and 14.86% (extracted with 0.1N citric acid) of dragon fruit peel pectin
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219 yield, respectively.

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220 Unlike WPD and WPF, there was a significant difference (p < 0.05) in pectin yield when
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221 extractions were carried out using dried and fresh forms of inner layer of dragon fruit peels (IPD

222 and IPF) (Table 2). The yield when IPF was used was the highest (26.38%) in comparison to

223 WPD, WPF, IPD and also to that found in previous studies on dragon fruit peel pectin. In

224 addition, the pectin yield found in this study (IPF) was the highest when compared to that of

225 mangosteen rind (12% using citrate phosphate buffer), passion fruit peel (14.80% using

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226 hydrochloric acid), apple pomace (14.55% using hydrochloric acid), lemon by-product (11.21%

227 using date juice), peach pomace (11.40% using 70% nitric acid), lemon peel (22-25% using

228 dilute mineral acids) and durian rind (7.1% using hydrochloric acid) as reported by Gan & Latiff

229 (2011), Kulkarni & Vijayanand (2010), Kumar & Chauhan (2010), Masmoudi et al. (2008),

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230 Pagan & Ibarz (1999), Seggiani et al. (2009) and Wai et al. (2010), respectively.

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231 3.2. Functional groups and degree of esterification (DE)

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232 Dragon fruit (DF) pectin extracted from WPD, WPF, IPD and IPF were abbreviated as

233 DF-WPD, DF-WPF, DF-IPD and DF-IPF, respectively. Preliminary characterisation of DF

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234 pectins was performed using FT-IR to investigate whether there is a difference between the
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235 samples. Fig. 2 shows the FT-IR spectra of the samples and citrus pectin. The spectra with the

236 wavenumber between 800 and 1,300 cm-1 is considered as the ‘finger print’ region for specific
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237 polysaccharide. It can be observed from the spectra that the samples have similar ‘finger print’
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238 region with that of citrus pectin suggesting that the extracted polysaccharide is from the pectin

family. A broader band of absorption around 3,400 cm-1 was attributed to the stretching of
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240 hydroxyl groups and an absorption band at 2,900 cm-1 was due to C–H stretching of CH2 groups
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241 (Tamaki et al., 2008).

242 Absorption bands appearing at 1,745 and 1,630 cm-1 were assigned to the stretching
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243 vibrations of ester carbonyl (C=O) groups and carboxyl ions (COO-), respectively (Singthong et
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244 al., 2004). A stronger absorption at 1,745 cm-1 coupled with a weaker absorption at 1,630 cm-1

245 indicated that DF-IPD and DF-IPF are the high methoxyl pectins (DE > 50%). A similar pattern

246 was observed for citrus pectin. However, the intensity of the absorption band at 1,745 cm-1 of

247 citrus pectin was stronger suggesting the DE of citrus pectin is probably higher than that of DF-

248 IPD and DF-IPF. It was noted that DF-WPD and DF-WPF have similar absorption intensity at

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249 the aforementioned two characteristic bands which needs further confirmation by calculating the

250 DE to predict whether they are high/low methoxyl pectin.

251 The intensity of the absorbance or band area of the ester carbonyl groups (1,745 cm-1)

252 increased and is the opposite for carboxylate stretching band (1,630 cm-1) with the increase in

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253 DE (Chatjigakis et al., 1998; Manrique & Lajolo, 2002; Singthong et al., 2004). The DE of DF-

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254 WPD, DF-WPF, DF-IPD, DF-IPF and citrus pectin determined using this FT-IR spectroscopic

255 technique were 53.74%, 51.71%, 62.88%, 63.74% and 68.00%, respectively. This finding further

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256 confirmed that all the samples and citrus pectin are high methoxyl pectins. It is worth

257 highlighting here that DF-IPF has the highest DE compared to other samples besides having the

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closest DE value to the commercial citrus pectin. Therefore, DF-IPF can be an alternative source
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259 of high methoxyl pectin.
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260 3.3. Proximate compositions of dragon fruit peel pectin

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261 Based on the yield and the preliminary characterisation using FT-IR, pectin from fresh
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262 inner layer of peel (DF-IPF) was used for subsequent physico-chemical characterisations as

263 follows. DF-IPF is identified as dragon fruit peel pectin henceforth. The proximate compositions
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264 of the pectin are shown in Table 3. Protein was detected in a small amount which will be usually

265 co-extracted during pectin extraction.

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266 3.4. Surface morphology of dragon fruit peel pectin

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267 The dragon fruit peel pectin appeared predominantly in large flakes with some blend of

268 irregular-shaped smaller flakes (Fig. 3). The smaller flakes might be other neutral

269 polysaccharides or proteins that were present in minor quantities as contaminants in the extracted

270 crude pectin since purification step was not carried out in this study.

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271 3.5. Homogeneity and molecular weight

272 The elution profiles of apple, citrus and dragon fruit peel pectins as a function of elution

273 volume obtained using HPSEC-MALLS-RI are illustrated in Fig. 4. RI gives a signal

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274 proportional to the concentration, whereas the MALLS response to a given concentration is

275 proportional to the product: concentration x molecular weight (Wyatt, 1993). The molecular

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276 characteristics of the pectins are shown in Table 4. The molecular weight (Mw) and chain length

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277 (Mn) of dragon fruit peel pectin was found to be lower than that of apple and citrus pectins.

278 Heterogenous profiles were observed, with dragon fruit peel pectin exhibiting a more

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279 pronounced sign of heterogeneity (Fig. 4c). A large polydispersity index (PDI) implies a wider
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280 molecular weight distribution and the PDI is equal to 1 for monodispersive polymer (Rogosic et

281 al., 1996). The larger PDI of dragon fruit peel pectin than that of apple and citrus pectins agreed
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282 well with the elution profile that is indicative of the higher heterogeneity of the sample.

283 A common peak eluted around 6.5 mL was detected in high intensity with the MALLS
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284 detector, which coincided with high RI intensity. Thus, a high molar weight component was
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285 present in a high concentration, and could be assigned to pectic polysaccharide. An intense RI

286 peak eluted around 8.5 mL (Fig. 4c), coincided with that with a minimal light scattering
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287 intensity, possibly due to the co-extracted high concentration of low molecular weight polymer
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288 together with the dragon fruit pectin which was absent in apple and citrus pectins. The co-eluted
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289 low molecular weight polymer with the predominant pectic polysaccharides could be the free

290 neutral polysaccharides or pectin side chains (Kravtchenko et al., 1992). A common peak eluted

291 around 9.5 mL could be attributed to the contaminant protein that is usually present in minor

292 amount in pectin. The root mean square radius of dragon fruit peel pectin was not significantly

293 different from that of the commercial pectins.

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294 3.6. Monosaccharide composition

295 Separation of ten derivatised monosaccharides is shown in Fig. 5. As expected, ribose,

296 glucuronic acid and fucose were not detected in the dragon fruit peel pectin. Xylose and

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297 arabinose were present in minor amounts (< 4%). Dragon fruit peel pectin was predominantly

298 constituted of galacturonic acid, followed by mannose, rhamnose, galactose and glucose (Table

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299 5). All the common monomers that are the building blocks of pectin were found in dragon fruit

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300 peel pectin in an acceptable ratio. Co-extraction of non-pectic substances such as traces of

301 heteromannans during pectin extraction would have contributed to a substantial amount of

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302 mannose in dragon fruit peel pectin. Significant rhamnose content in the peel could be attributed
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303 to the presence of rhamnogalacturonan I region apart from the predominant co-polymer block of

304 complex pectin, the unsubstituted homogalacturonans (Vincken et al., 2003; Yapo, 2011). The
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305 presence of other neutral sugars such as galactose and arabinose in the peel suggest the possible

306 presence of galactan and arabinan as side chains of rhamnogalacturonan I as documented by

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307 Hokputsa et al. (2004), Kurita et al. (2008) and Prabasari et al. (2011) with pectins extracted
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308 from durian rind, citrus peel and orange albedo, respectively.
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309 3.7. Flow behaviour of dragon fruit peel pectin

310 Fig. 6 presents the flow behaviour of dragon fruit peel pectin solution in comparison to
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311 that of apple and citrus pectin solutions. At concentrations 0.5% and 1.0%, a linear relationship
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312 between the shear stress and shear rate was observed in the shear rate range of 0 - 400 s-1 (Fig. 6a

313 and 6b) that showed the Newtonian behaviour of pectin at low concentrations. At concentration

314 0.5%, Yaseen et al. (2005) also found that pectin demonstrated a Newtonian behaviour. Marcotte

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315 et al. (2001) reported that 1% pectin exhibited a nearly Newtonian like flow behaviour (flow

316 behaviour index = 0.96, close to 1).

317 As expected, the viscosity of pectin solutions increased with an increase in pectin

318 concentration. Pseudoplastic behaviour of pectin was observed at higher concentrations (Fig. 6c

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319 and 6d). An increase in pseudoplasticity with an increase in pectin concentration was also

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320 noticed by Iagher et al. (2002), Marcotte et al. (2001) and Vardhanabhuti & Ikeda (2006). This

321 phenomenon is common for polysaccharides where the zero shear viscosity value becomes

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322 higher with the increase in polymer concentration. At the same time the Newtonian plateau limit

323 is shifted to a low shear region (Iagher et al., 2002).

324
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Dragon fruit peel pectin demonstrated similar viscosity profile to that of citrus pectin but
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325 lower than that of apple pectin (Fig. 6). The low viscosity of dragon fruit peel pectin can be

attributed to its lower Mw and Mn. This suggests that it can be added to low viscous foods and
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326

327 beverages as a stabiliser or thickener. It might even be a potential wall material for spray drying
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328 of juices due to its shorter chains and low viscosity. Preliminary work in our laboratory also
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329 showed that it can be utilised as a gelling agent in jams.

330 4. Conclusions
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331 High methoxyl pectin was extracted from dragon fruit peel using 1% citric acid. The
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332 physico-chemical characteristics of the dragon fruit peel pectin were comparable to that of
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333 commercial pectins except for the viscosity. However, the dragon fruit peel pectin can be utilised

334 as a functional ingredient in low viscous foods. Both technological and health functional

335 characterisations should be performed to further understand its behaviour in food and the health

336 benefits it may provide.

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337 Acknowledgement

338 We gratefully acknowledge the financial support received from Universiti Putra Malaysia

339 Research Grant No. 02-01-11-1139 RU.

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340 References

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462

463

464

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465 Figure captions

466 Fig. 1. Response surface plot showing the significant (p < 0.05) interaction effect of time (min)
467 and pH on fresh inner layer of peel (IPF).pectin yield (%).
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469 Fig. 2. FT-IR spectra of DF-WPD, DF-WPF, DF-IPD, DF-IPF and citrus pectins. DF: dragon
470 fruit; WPD: whole peel (dried); WPF: whole peel (fresh); IPD: inner layer of peel (dried); IPF:

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471 inner layer of peel (fresh).
472
473 Fig. 3. Surface morphology of dragon fruit peel pectin at (a) x 25 and (b) x100 magnifications.

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474 Fig. 4. Elution profiles obtained by HPSEC-MALLS-RI using (a) apple pectin, (b) citrus pectin
475 and (c) dragon fruit peel pectin.

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476 Fig. 5. Chromatogram of derivatised monosaccharide standards and dragon fruit peel pectin.

477 Fig. 6. Flow curves of apple, citrus and dragon fruit peel pectins at concentrations (a) 0.5%, (b)

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478 1.0%, (c) 2.0% and (d) 3.0%.
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613 Table 1
614 The matrix of central composite design and the experimental data obtained for the response
615 variable (yield, %) using the fresh inner layer of peel (IPF).
Run Order Blocks Temperature Time pH Yield
(oC) (min) (%)
1* 3 67.5 95 3.25 13.9

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2 3 67.5 95 4.47 4.9
3 3 38.9 95 3.25 9.3
4 3 96.0 95 3.25 18.8

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5 3 73.0 67 2.03 26.4
6 3 67.5 185 3.25 13.4
7* 3 67.5 95 3.25 14.2

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8 3 67.5 5.2 3.25 7.2
9* 2 67.5 95 3.25 14.5
10 2 50.0 150 2.50 23.3
11* 2 67.5 95 3.25 13.2

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12 2 85.0 150 4.00 6.0
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13 2 50.0 40 4.00 5.6
14 2 85.0 40 2.50 21.8
15* 1 67.5 95 3.25 14.0
16 1 85.0 40 4.00 6.8
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17 1 50.0 150 4.00 6.4

18 1 50.0 40 2.50 20.7
19 1 85.0 150 2.50 24.5
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20* 1 67.5 95 3.25 14.3

*
616 Centre point.
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634 Table 2
635 Moisture content (%) of dragon fruit peel fractions and the pectin yield (%).
Pectin yield
Samplea Moisture (%) (% dry weight)
WPD b
7.15 ± 0.07 15.00b ± 0.48
WPF 89.94c ± 0.12 14.41b ± 0.79
7.58d ± 0.08 7.99c ± 0.92

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IPD
IPF 91.68e ± 0.07 26.38d ± 0.47
636 Data are expressed as means ± standard deviations of triplicate analysis.
a
WPD: whole peel (dried); WPF: whole peel (fresh); IPD: inner layer of peel (dried); IPF: inner

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638 layer of peel (fresh).
b-e
639 Means followed by different superscript lowercase letters indicate significant differences (p <
0.05) within column by Tukey’s HSD test.

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672 Table 3
673 Proximate compositions of dragon fruit peel pectin based on dry weight basis (g/100g).
Composition (%) Dragon fruit peel pectin
Moisture 7.81 ± 0.10
Ash 4.73 ± 0.09
Total fat 0.04 ± 0.01
Protein 1.15 ± 0.03

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Dietary fibre 62.76 ± 0.05
Others (by difference) 23.51 ± 0.06
674 Data are expressed as means ± standard deviations of triplicate analysis.

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710 Table 4
711 Molecular characteristics of apple, citrus and dragon fruit peel pectins.
Dragon fruit peel
Molecular parametera Apple pectin Citrus pectin
pectin
Mw (x 105 Da) 1.44b ± 0.10 1.38b ± 0.01 0.88c ± 0.06
5 b
Mn (x 10 Da) 0.65 ± 0.04 0.64b ± 0.03 0.11c ± 0.01
Polydispersity index (Mw/Mn) 2.22b ± 0.20 2.17b ± 0.09 8.12c ± 1.55

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b
RMS radius (nm) 35.37 ± 0.91 35.03b ± 0.45 35.00b ± 1.45
712 Data are expressed as means ± standard deviations of triplicate analysis.
a
713 Mw: weight average molar mass; Mn: number average molar mass; RMS: root mean square

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b-c
714 Means followed by different superscript lowercase letters indicate significant differences (p <
715 0.05) within row by Tukey’s HSD test.

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749 Table 5
750 Monosaccharide composition (%) of dragon fruit peel pectin.
Monosaccharide Composition (%)
Mannose 17.78 ± 1.07
Ribose n.d.
Rhamnose 14.47 ± 0.39
Glucuronic acid n.d.

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Galacturonic acid 39.11 ± 1.87
Glucose 10.82 ± 0.44
Galactose 11.91 ± 0.48

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Xylose 2.41 ± 0.02
Arabinose 3.49 ± 0.07
Fucose n.d.

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751 Data are expressed as means ± standard deviations of triplicate analysis. n.d. : not detected.
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791 Figure 1

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793 Hold values:

Temperature
67.5°C
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817 Figure 2

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DF-WPD
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DF-WPF

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DF-IPD

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DF-IPF

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826 Citrus pectin

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843 Figure 3

844 a
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869 Figure 4

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896 Figure 5

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898 Standard
Dragon fruit peel pectin
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922 Figure 6

923 a

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Highlights

• The yield of pectin extracted from red dragon fruit peel was higher than that reported for
other fruit peels.

• The low molecular weight and viscosity of the pectin allow its use as wall material for
spray drying of juices.

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• The pectin is more heterologous than other pectins indicating its functional versatility.

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