Oligosaccharides of pitaya (dragon fruit) esh and their prebiotic properties

S. Wichienchot
a,
*
, M. Jatupornpipat
b
, R.A. Rastall
c
a
Nutraceutical and Functional Food Research and Development Center, Faculty of Agro-Industry, Prince of Songkla University, Hat Yai, Songkhla 90112, Thailand
b
Department of Applied Biology, Faculty of Science, King Mongkuts Institute of Technology Ladkrabang, Bangkok 10520, Thailand
c
School of Food Biosciences, The University of Reading, Whiteknights, Reading, UK
a r t i c l e i n f o
Article history:
Received 26 May 2009
Received in revised form 2 November 2009
Accepted 9 November 2009
Keywords:
Pitaya
Dragon fruit
Oligosaccharides
Prebiotic
Probiotic
a b s t r a c t
The major carbohydrates of white and red-esh pitayas (dragon fruit) were glucose, fructose and some
oligosaccharides (total concentrations of 86.2 and 89.6 g/kg, respectively). The molecular weight distribu-
tion of the extract was affected by the extraction solvent. The maximum oligosaccharides content
(27.40%), which included fractions with molecular weights of 273275, 448500 and 787911 Da, were
obtained using 80% ethanol extraction at room temperature (28 2 C). The low molecular weight frac-
tion, including glucose and fructose, was successfully removed by yeast cultivation. The molecular
weights of mixed oligosaccharides (716, 700, 490 and 474 Da) were conrmed by mass spectrometry.
The mixed oligosaccharides showed that they were resistant to hydrolysis by articial human gastric
juice and human a-amylase, giving maximum hydrolysis of 4.04% and 34.88%, respectively. The mixed
oligosaccharides were also found capable of stimulating the growth of lactobacilli and bidobacteria.
2009 Elsevier Ltd. All rights reserved.
1. Introduction
Pitaya (Hylocereus undatus (Haw.)) or dragon fruit is native to
Mexico, Central and South America (Benzing, 1990; Haber, 1983)
and has been grown in Vietnam for at least 100 years, following
its introduction by the French (Mizrahi, Nerd, & Nobel, 1997). A
member of the Cactaceae, it has trailing cladode stems modied
to act as leaves and bears spectacular ovoid fruit year-round which
has a bright red colour when mature, and contains white, crimson,
or pale yellow-esh, depending on the cultivar, interspersed with
small black seeds. In Thailand, one of the widely grown varieties
is H. undatus or red pitaya with white-esh. Other varieties that
have been commercialised are Hylocereus polyrhizus (red pitaya
with red-esh) and Hylocereus megalanthus (yellow pitaya) (Bar-
beau, 1990). Currently, there is much interest in developing this
crop for fresh fruit export beyond the local Asian markets of Sin-
gapore, Hong Kong, Taiwan, Philippines, Malaysia and Thailand
(Hoa, Clark, Waddell, & Woolf, 2006; Mizrahi et al., 1997).
Dragon fruit has been reported as a source of beta-carotene,
lycopene and vitamin E, with average concentrations of 1.4, 3.4
and 0.26 lg/100 g edible portion, respectively (Charoensiri, Kon-
gkachuicha, Suknicom, & Sungpuag, 2009). The seed of dragon fruit
contains 50% essential fatty acids, i.e., 48% linoleic acid (C18:2) and
1.5% linolenic acid (C18:3) (Arifn et al., 2008). Thus, dragon fruit
has potential for use as a source of functional ingredients to pro-
vide nutrients that may prevent nutrition-related diseases and im-
prove physical and mental well-being of the consumers.
Prebiotics are non-digestible oligosaccharides that benecially
affect the host by stimulating the growth and/or activity of one
or a limited number of bacteria in the colon, thus improving host
health (Gibson & Roberfroid, 1995). Ten samples from thirteen
fruits and vegetables and their parts from southern Thailand were
reported as potential sources of natural prebiotics with the highest
oligosaccharide content being 9.81% (w/w) (Thammarutwasik
et al., 2009). Dragon fruit was not included in that study. Chicory
and artichoke root have been used as natural sources of inulin
and oligofructose in the commercial production of prebiotics, hav-
ing oligosaccharide yields in the range of 1820% (Gibson & Rastall,
2006). As a potential source of high-yielding oligosaccharide for
commercial prebiotic production, dragon fruit was selected for
investigation, with respect to optimal extraction method, sugar
content and prebiotic properties.
2. Materials and methods
2.1. Samples, chemicals, media and microorganisms
White-esh dragon fruits ( H. undatus) were imported from
Hoang Hau Dragon Fruit Farm, Co. Ltd., Vietnam. The fruits were
mediumsize packed in a padded paper box. Red-esh dragon fruits
(H. polyrhizus) were obtained from a local market in Chumphon,
Thailand. Unless otherwise stated, all chemical reagents and en-
zyme were obtained from SigmaAldrich Co. Ltd. Cheesecloth bags
0308-8146/$ - see front matter 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.foodchem.2009.11.026
* Corresponding author. Tel.: +66 74286389; fax: +66 74212889.
E-mail addresses: santad.w@psu.ac.th, wsantad@yahoo.com (S. Wichienchot).
Food Chemistry 120 (2010) 850857
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were prepared (23 14 cm). Anaerobic gas pack and Reinforced
Clostridium Medium were supplied by Oxoid, Ltd., Basingstoke,
UK. MRS broth was purchased from Merck, Co., Ltd., Darmstadt,
Germany and Rogosa SL Agar was purchased from Difco, Co., Ltd.,
USA. Bidobacterium bidum NCIMB 702715 was obtained from
the National Collections of Industrial Food and Marine Bacteria,
Aberdeen, UK. Lactobacillus delbrueckii BCC 13296 and Saccharomy-
ces cerevisiae BCC 12652 were obtained from BIOTEC Culture Col-
lection, Bangkok, Thailand.
2.2. Determination of sugar composition
Reducing sugar concentration was determined using the
dinitrosalicylic acid (DNS) assay and expressed as maltose equiva-
lents (Robertson et al., 2001). The assay was calibrated with malt-
ose standards from 0 to 1 mg/ml. Sample or glucose standard
solution (1 ml) was added to DNS solution (1 ml) and water
(1 ml). The mixture was boiled for 15 min and after cooling on
ice for 2 min, water (9 ml) was added before mixing and measuring
the absorbance at 540 nm. Content of reducing sugar in the sample
was compared to the standard curve of glucose. The total sugar
concentration, expressed as glucose equivalents, was determined
using the phenolsulphuric acid assay (Dubois, Gilles, Hamilton,
Rebers, & Smith, 1956). The assay was calibrated with D-glucose
standards from 0 to 100 lg/ml. Sample or glucose standard solu-
tion (1 ml) was added to 5% (w/v) phenol solution (1 ml). Concen-
trated sulphuric acid (5 ml) was then added and left for 10 min
prior to vigorous mixing. The samples and standards were left for
20 min at room temperature before reading the absorbance at
490 nm. The concentration of carbohydrate in the sample was cal-
culated by comparison with the standard curve of glucose. Mono-
saccharide and disaccharide were analysed by HPLC. Sample was
prepared in solution (1% w/v) and ltered through 0.45-lm
membrane; 20 ll sample were injected in a Zorbax Carbohydrate
column (4.6 150 mm, 5 lm resin). Mobile phase was acetoni-
trile:water (75:25) and ow rate was 0.5 ml/min. Refractive Index
(RI) detector was used and column oven temperature was 35 C.
Glucose and fructose were used as monosaccharide standards
and sucrose was used as a disaccharide standard. Qualitative anal-
ysis of sugar in the sample was determined by comparison to the
retention time of standard sugar. The concentration of sugar in
the sample was calculated by comparison with peak area of the
standard curve of the respective sugar.
2.3. Determination of molecular weight distribution of
oligosaccharides
Molecular weight of sample was analysed by gel permeation
chromatography (GPC) with refractive index detection. The analyt-
ical system consisted of Ultrahydrogel linear column (MWCO
100020,000,000) and Ultrahydrogel 120 column (MWCO 100
5000) connected in series with decreasing pore size. The ow rate
was 0.6 ml/min, the eluent was 0.1 M sodium nitrate and the oper-
ating temperature was 30 C. Samples and standards (pullulan
with different molecular weight, maltoheptaose, and glucose) were
ltered through a 0.45-lm syringe lter and the injection volume
was 20 ll. The system was calibrated with 8 standards consisting
of pullulan with average molecular weights of 788, 404, 112,
47.3, 22.8, 5.9 kDa, maltopentaose (0.828 kDa) and glucose
(0.180 kDa). External standards of maltopentaose (0.828 kDa) at
concentrations of 110 mg/ml were used to establish the area re-
sponse for quantication of the products. Oligosaccharides yield
was calculated by the summary of the percentage area of the oligo-
saccharides fraction. The molecular weight (MW) of oligosaccha-
rides was determined by comparing sample retention time to
those of the standard curve (Mountzouris, Gilmour, Jay, & Rastall,
1999). The molecular weight distribution of mixed oligosaccha-
rides after removal of low molecular weight fraction by yeast cul-
tivation was conrmed by mass spectrometry (Micromass, UK)
with ESI detector in positive mode (modied from Hernandez,
Muiz-Matute, Olano, Moreno, & Sanz, 2009).
2.4. Purication method
The extract obtained by 80% ethanol extraction at ambient tem-
perature was distilled by rotary evaporator at 175 mbar, 60 C and
rotating speed of 45 rpm. The concentrated extract was diluted
with distilled water in the ratio of 1:1 and redistilled again. This
step was repeated twice to removal residual ethanol in the extract.
Ethanol-free extract was adjusted to 20 Brix by addition of dis-
tilled water before 5% (v/v) inoculum (18 h culture of S. cerevisiae
BCC 12652) was inoculated. Cultivation was carried out at room
temperature (28 2 C) for 48 h to remove mostly glucose and
some fructose. Culture broth was then diluted with sterile distilled
water to 10 Brix and cultivation was carried out under the same
conditions for 6 h to remove mostly the remaining fructose in
the crude extract (modied from Hernandez et al., 2009).
2.5. Effect of extraction methods
White-esh dragon fruits ( H. undatus) and red-esh dragon
fruits ( H. polyrhizus) were used as raw material for this study.
The dragon fruit was peeled and the esh was extracted with 3 dif-
ferent solvents (distilled water, 20% ethanol and 80% ethanol).
Extraction by water at room temperature was done by mixing
water and the esh in the ratio of 2:1, stirring continuously and
leaving for 1 h at room temperature. The mixture was then ltered
through a cheesecloth bag while being pressed with a paddle blen-
der (Seward-Stomacher 400) to remove the seeds. Viscous juice ob-
tained was further evaporated by a rotary evaporator under
reduced pressure at 40 C. Sample was dried with a freeze dryer
before sugar composition, molecular weight distribution and pre-
biotic properties were analysed. Hot water extraction was done
by mixing hot water and the esh in the ratio of 2:1, stirring con-
tinuously and leaving for 1 h in a controlled temperature water
bath (85 2 C). The mixture was then ltered through a cheese-
cloth bag while being pressed by a stomacher to remove the seeds.
The sample was then dried and further analysed as mentioned
above. Extraction by 20% and 80% (v/v) ethanol were done using
the same manner and the mixture was left for 1 h at room temper-
ature. The sample was then dried and again analysed as above.
2.6. Effect of articial human gastric juice hydrolysis
Mixed oligosaccharides obtained by 80% ethanol extraction and
the juice without extraction from white-esh were tested for acid
resistance, compared to an inulin standard as a commercial prebi-
otic reference. Sample was dissolved in RO water to give a 1% (w/v)
solution. Articial human gastric juice was mimicked by using
hydrochloric acid buffer containing (in g/l): NaCl, 8; KCl, 0.2; Na
2
H-
PO
4
2H
2
O, 8.25; NaHPO
4
, 14.35; CaCl
2
2H
2
O, 0.1; MgCl
2
6H
2
O, 0.18.
The pH of the buffer was adjusted to 1, 2, 3, 4 and 5 using 5 M HCl
(Korakli, Ganzle, & Vogel, 2002). HCl buffer (5 ml) at each pH was
added to the sample solution (5 ml) and the reaction mixture
was incubated in a water bath at a controlled temperature of
37 1 C for 6 h. Sample (1 ml) was taken periodically at 0, 0.5, 1,
2, 4 and 6 h. Reducing sugar content in the sample was determined
by DNS method (Robertson et al., 2001) and total sugar was deter-
mined by phenolsulphuric acid method (Dubois et al., 1956). Per-
centage hydrolysis of sample was calculated based on reducing
sugar liberated and total sugar content of the sample (Korakli
et al., 2002):
S. Wichienchot et al. / Food Chemistry 120 (2010) 850857 851
Hydrolysis%

reducing sugar released 100

total sugar content initial reducing sugar content
1
where reducing sugar released is the difference between its nal
and initial content.
2.7. Effect of human a-amylase hydrolysis
The enzyme activity was determined by the Sigma quality con-
trol test procedure for a-amylase (EC. 3.2.1.1). Enzyme was pre-
pared in solution containing 2 units/ml using sodium phosphate
buffer (20 mM) in sodium chloride (6.7 mM), pH 4, 5, 6, 7 or 8.
Sample was prepared as a 1% (w/v) solution in sodium phosphate
buffer. Five millilitres of enzyme solution were added to 5 ml of
sample solution. The reaction mixture was incubated in a con-
trolled temperature water bath (37 1 C) for 6 h. Sample (1 ml)
was taken at 0, 0.5, 1, 2, 4 and 6 h to determine reducing sugar
and total sugar. Percentage hydrolysis was calculated based on
reducing sugar liberated and the total sugar content of the sample
as above.
2.8. Effect of probiotic growth
Mixed oligosaccharides obtained from white-esh and a refer-
ence prebiotic (inulin) were used as carbon source for the growth
of 2 probiotic strains. Inoculumof L. delbrueckii BCC 13296 was pre-
pared by cultivation on MRS broth at 37 C for 24 h. One millilitre
of sample solution was prepared (1% w/v) and added to MRS broth
(3 ml), then the inoculum (1 ml) was added, to obtain a nal con-
centration of 10
8
cell/ml. The culture was incubated at 37 C for
48 h in anaerobic conditions. Sample was taken at 0, 24 and 48 h
for bacterial enumeration by standard direct count using a haem-
ocytometer (Swanson, Petran, & Hanlin, 2001). B. bidum 702715
was sub-cultured in Reinforced Clostridial Medium at 37 C for
24 h prior to use as inoculum. One millilitre of sample solution
was prepared (1% w/v) and aseptically added to Reinforced Clos-
tridial Medium (3 ml) in a sterilised screwed-cap vial with rubber
septum. Cultivation was carried out under anaerobic conditions, by
ushing with nitrogen gas over the culture medium then closing
with a screwed-cap. Inoculum (1 ml) was injected by a sterilised
syringe into the sealed vial via a septum. Cultivation was carried
Fig. 1. Sugar composition of white-esh (a) and red-esh (b) dragon fruit extracts, analysed by HPLC.
Table 1
Physical and chemical properties of two varieties of pitaya.
Characteristic White-esh dragon fruit Red-esh dragon fruit
Fruit dimension (cm)
a
Length 134 5.0a 127 5.5b
Diameter 94.0 9.0a 66.0 4.0b
Fruit weight (g)
a
Flesh weight 305 75.0a 215 35.0b
Skin weight 100 30.0a 75.0 25.0b
Sweetness (Brix)
a
12.5 0.55a 14.8 0.75b
Sugar content (g/kg)
b
Glucose 353 0.74a 401 1.27b
Fructose 238 0.84a 158 0.32b
Oligosaccharides 86.2 0.93a 89.6 0.76a
Different letters in the same row mean a signicant difference (p 6 0.05).
a
Average of 15 fruits.
b
Average of triplicate analyses by HPLC.
852 S. Wichienchot et al. / Food Chemistry 120 (2010) 850857
out under anaerobic conditions at 37 C for 72 h. Sample was taken
at 0, 24, 48 and 72 h for bacterial enumeration by a standard direct
count using haemocytometer (Swanson et al., 2001).
3. Results and discussion
3.1. Chemical composition and molecular weight distribution of
oligosaccharides
Sugars of white- and red-esh dragon fruit analysed by HPLC
consisted mostly of glucose, fructose and some oligosaccharides,
as shown in Fig. 1. Glucose concentration of white-esh
(353 0.7 g/kg) was signicantly lower than of red eshed fruit
(401 1.27 g/kg) (p 6 0.05). In contrast, fructose of white-esh
(238 0.84 g/kg) was signicantly higher than red-esh (158
0.32 g/kg) (p 6 0.05), as shown in Table 1. Oligosaccharides content
in red-esh (89.6 0.76 g/kg) was slightly higher than white-esh
(86.2 0.93 g/kg) but not signicantly. Molecular weight distribu-
tions of crude extract obtained from either white or red-esh ana-
lysed by gel permeation chromatography showed exactly the same
values (273275, 448500 and 787911 Da by 80% ethanol extrac-
tion and 273275 and 787911 Da by distilled water extraction,
Fig. 2). The low molecular weight fraction (273275 Da), glucose
and fructose were successfully removed by two-step yeast
(S. cerevisiae) cultivation and molecular weight distribution of the
Fig. 2. Molecular weight distribution of white-esh dragon fruit extracts obtained by water (a) and 80% ethanol (b) extraction at ambient temperature, analysed by GPC.
S. Wichienchot et al. / Food Chemistry 120 (2010) 850857 853
mixed oligosaccharides was conrmed by mass spectrometry. The
mixed oligosaccharides consisted of four components of 716, 700,
490 and 474 Da with relative percentages of 100, 68, 45 and 21,
respectively (Fig. 3). Thus, the degree of polymerisation of mixed
oligosaccharides was 34, which was in the same range as some
fructooligosaccharides, i.e., 1-kestose, 6-kestose and neokestose
(1 glucose unit and 2 fructose units), or nystose, bifurcose and neo-
bifurcose (1 glucose unit and 3 fructose units), or stachyose (3 glu-
cose units and 1 fructose unit). Some plants producing prebiotic
oligosaccharides, such as onion, garlic, dahlia, tulip, oat (Ritsema
& Smeekensy, 2003), soy bean and mung bean, contained oligosac-
charides in the same range of molecular weight as this study;
chicory and artichoke contained higher molecular weight oligosac-
charides (Eggleston & Cote, 2003; Xiaoli et al., 2008). The molecular
weight of the pitaya oligosaccharides was in the same range as
some commercial prebiotics, such as fructooligosaccharide syn-
thesised from sucrose (DP = 24), oligofructose (DP = 4) derived
from enzymatic hydrolysis of inulin, and soybean oligosaccharide
obtained from soybean extraction (DP = 45) (Vernazza, Rabiu, &
Gibson, 2006).
3.2. Effect of extraction methods
Though white- and red-esh dragon fruit were not signicantly
different in terms of quality and quantity of their oligosaccharides,
white-esh variety was less expensive and more abundant in Thai-
land, and it was thus chosen for subsequent studies. For the study
on extraction methods, it was found that solvent had marked ef-
fects on the molecular weight of the oligosaccharides obtained. Oli-
gosaccharides obtained by water extraction, either at ambient
temperature (28 2 C) or 85 2 C had molecular weights of
787911 Da while with 80% ethanol extraction, a fraction of
448500 Da was also found (Fig. 2). The yield of oligosaccharides
obtained by water extraction at ambient temperature was 15.1%
while 80% ethanol extraction elevated the yield to 27.4% (Table
2). Thus extraction methods had a highly signicant effect
(p 6 0.01) on the oligosaccharide yield obtained. It was concluded
that the optimal extraction method for oligosaccharides from
white-esh was 80% (w/v) ethanol (ethanol:esh was 2:1, v/v) at
ambient temperature. To obtain higher purity of pitaya oligosac-
charides, a further purication step is required, i.e., chromatogra-
phy or membrane ltration, which has been used for commercial
prebiotic production (Kono, 1993), whereas in this work purica-
tion was done with a biological approach using yeast. The results
showed that S. cerevisiae was very efcient at removing mostly glu-
cose and some of fructose in the crude extract within 48 h whereas
removal of residual fructose required 6 h after the extract was
diluted twice (data not shown). However, this method was less
efcient for the removal of lactose and galactose from galactooligo-
saccharide mixtures (Hernandez et al., 2009). This is because the
native strain of S. cerevisiae could metabolize common sugars such
as glucose, fructose, sucrose and maltose, but not lactose, melibi-
ose and xylose as a carbon source.
3.3. Effect of articial human gastric juice hydrolysis
Mixed oligosaccharides obtained with 80% ethanol extraction
from white-esh were hydrolysed with articial human gastric
juice and showed to be resistant to the juice. Percentage of hydro-
lysis increased with decreasing pH of articial gastric juice (Fig. 4).
The degree of hydrolysis at pH of 1, 2, 3, 4 and 5 was 4.07%, 2.43%,
1.66%, 0.85% and 0.02%, respectively. The maximum hydrolysis
(4.07%) occurred after 2 h of incubation at pH 1. Pitaya oligosaccha-
rides showed high acid resistance compared to the reference prebi-
otic (inulin), which gave maximum hydrolysis of 55.00%, 23.22%,
35.75% and 5.96% at pH 1, 2, 3, 4 and 5, respectively. Food is usually
retained in the human stomach where gastric juice (pH 24) is
released within 2 h; thus, when pitaya oligosaccharides are
Fig. 3. Molecular weight distribution of white-esh dragon fruit extract after removal of low molecular weight sugar by yeast cultivation, analysed by mass spectrometry.
854 S. Wichienchot et al. / Food Chemistry 120 (2010) 850857
consumed, 96% of them is estimated to reach the intestine. This re-
sult is comparable to that of kojiooligosaccharide, which had 100%
resistance to articial gastric acid (Nakada, Nishimoto, Chaen, &
Fukuda, 2003). Gluco-oligosaccharide produced by Gluconobacter
oxydans NCIMB 4943 showed 98.4% resistant in the acidic condi-
tions of the human stomach (Wichienchot, Prasertsan, Hongpatta-
rakere, Gibson, & Rastall, 2006). Dextran has been reported to be
highly resistant to acidic conditions in the human digestive tract
(Debnam, Denholm, & Grimble, 1998).
3.4. Effect of human a-amylase hydrolysis
The hydrolysis of oligosaccharides from white-esh dragon fruit
with human salivary a-amylase at pH 48 showed that higher pH
gave a signicantly higher degree of hydrolysis (p 6 0.05); how-
ever, there was no signicant difference at pH 4 and 5. The maxi-
mum hydrolysis at pH 4, 5, 6, 7 and 8 was 2.90%, 3.08%, 3.28%,
6.88% and 11.18%, respectively after 30 min incubation (Fig. 5).
The maximum hydrolysis of a reference prebiotic (inluin) in-
creased at higher pH, giving values of 5.72%, 8.03%, 7.39%, 12.32%
and 16.22% at pH 4, 5, 6, 7 and 8, respectively. Thus, pitaya
oligosaccharides had signicantly higher enzymatic resistance
compared to inulin (p 6 0.01). It was estimated that about 50% of
the pitaya oligosaccharides consumed would reach the colon since
some of them were hydrolysed by a-amylase (16%), by acid in
stomach (2.5%) and by brush-border enzymes in small intestine
(30%). Typically, the carbohydrates were mainly digested in the
small intestine (30%) where some brush-border enzymes, i.e.,
isomaltase, glucoamylase, maltase, sucrase and lactase, hydrolyse
Table 2
Sugar contents of white-esh dragon fruit obtained by different extraction methods.
Sugar content
a
Extraction by water Extraction by ethanol
Ambient Hot 20% (w/v) 80% (w/v)
Mono- and disaccharide (%) 84.9 1.8a 83.6 2.1a 79.4 2.8b 72.6 1.6c
Oligosaccharides (%) 15.1 0.5a 16.4 0.6a 20.6 1.3b 27.4 1.5c
Different letters in the same row mean a signicant difference (p 6 0.05).
a
% area of fractions analysed by GPC in triplicate.
0.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
Time (h)
H
y
d
r
o
l
y
s
i
s

(
%
)
0
50
100
150
200
250
300
350
400
450
0 1 2 3 4 5 6
0 1 2 3 4 5 6
Time (h)
H
y
d
r
o
l
y
s
i
s

(
%
)
a
b
Fig. 4. Resistance of white-esh dragon fruit oligosaccharides (a) and inulin (b) to
various articial gastric juices; incubation for 6 h at 37 C. , pH 1; j, pH 2; N, pH 3;
, pH 4; s, pH 5.
0
5
10
15
20
25
30
35
40
Time (h)
H
y
d
r
o
l
y
s
i
s

(
%
)
0
5
10
15
20
25
30
35
40
45
50
0 1 2 3 4 5 6
0 1 2 3 4 5 6
Time (h)
H
y
d
r
o
l
y
s
i
s

(
%
)
a
b
Fig. 5. Resistance of white-esh dragon fruit oligosaccharides (a) and inulin (b) to
human salivary a-amylase (1 U/ml) at various pH values, incubated at 37 C for 6 h.
, pH 4; j, pH 5; N, pH 6; , pH 7; s, pH 8.
Table 3
Microbial counts of Lactobacillus delbrueckii BCC 13296 using extracts obtained from
white-esh dragon fruit and inulin as carbon sources.
Sample Microbial counts (cell/ml)
0 h 24 h 48 h
Crude extract 9.72 10
7
a 6.31 10
8
a 2.55 10
9
a
Oligosaccharides extract 9.02 10
7
a 6.42 10
8
a 6.17 10
9
b
Inulin 9.14 10
7
a 7.47 10
8
b 8.24 10
8
c
Average of triplicate analyses.
Different letters in the same column mean a signicant difference (p 6 0.05).
S. Wichienchot et al. / Food Chemistry 120 (2010) 850857 855
a-1,4- and a-1,6-linked glucosaccharides present in the small
intestine and yield monosaccharides as end-products (Johnson &
Schmit, 2005). It has been reported that 88% of inulin and oligo-
fructose reach the colon, using both the ileostomy patient model
and the intubation model (Cummings & Macfarlane, 2002). It has
also been reported that, using an in vitro technique, at least 60%
of gluco-oligosaccharide produced by G. oxydans NCIMB 4943
reaches the colon. Therefore, the oligosaccharides in this experi-
ment appeared to have partial enzymatic resistance.
3.5. Effect of pitaya oligosaccharides on probiotic growth
Oligosaccharides extracted from white-esh dragon fruit were
used as a carbon source for the cultivation of two probiotic strains.
It was found that the oligosaccharides stimulated the growth of L.
delbrueckii BCC13296 by increasing its number from 9.02 10
7
to
6.17 10
9
cell/ml within 48 h. Although inulin also stimulated
the growth of this probiotic strain, it had a lower effect (Table 3).
It has been reported that oligosaccharides extracted from jackfruit
seed also stimulated the growth of lactobacilli (Thammarutwasik
et al., 2009). Pitaya oligosaccharides also stimulated the growth of
B. bidum NCIMB 702715 by increasing their number from
1.70 10
8
to 2.51 10
9
cell/ml within 72 h (Table 4); however,
inulin showed a greater effect but no signicant difference. Jackfruit
seed oligosaccharides have been reported to stimulate the growth
of lactobacilli and bidobacterium (Thammarutwasik et al., 2009).
A similar result was observed on gluco-oligosaccharide produced
by G. oxydans (Wichienchot, Prasertsan, Hongpattarakere, Rastall,
& Gibson, 2006). Some commercial prebiotics that have been
reported to stimulate the growth of bidobacterium were inulin,
oligofructose, fructooligosaccharide, galactooligosaccharide, genti-
ooligosaccharide, isomaltooligosaccharide, lactosucrose, lactulose,
rafnose, soybean oligosaccharide and xylooligosaccharide (Sako,
Matsumoto, & Tanaka, 1999). Pitaya oligosaccharides showed other
functional properties beside their prebiotic effect, such as reduced
caloric intake and insulinaemia, compared to digestible carbohy-
drates. Therefore, oligosaccharides from dragon fruit may be suit-
able for inclusion as food supplements in a wide variety of food
products, e.g., dairy products, products designed for overweight
individuals, diabetic prevention products and prebiotic products.
4. Conclusion
Oligosaccharide content in the white- and red-esh pitaya was
86.2 and 89.6 g/kg, respectively. The optimal extraction conditions
for pitaya esh were 80% (w/v) ethanol, with a ratio of solvent to
esh of 2:1 at ambient temperature (28 2 C). The oligosaccha-
ride content in the crude extract was 27.40% relative to the sugar
content. Low molecular weight fraction including glucose and fruc-
tose in the crude extract was successful removed by yeast (S. cere-
visiae) cultivation. Partial puried extract consisted of a mixture of
oligosaccharides having molecular weights of 716, 700, 490 and
474 Da with a degree of polymerisation of 34. Pitaya oligosaccha-
rides showed prebiotic properties, which included resistance to
acid conditions in human stomach, partial resistance to human sal-
ivary a-amylase and the capability to stimulate the growth of lac-
tobacilli and bidobacteria. Thus, pitaya is a potential source of
prebiotics, which may be used as an ingredient in functional food
and nutraceutical products.
Conict of interest statement
The work described in the manuscript is original. It has never
been presented, submitted or published in any publication before
and does not encroach on any patents.
Statement of authorship
S. Wichienchot participated in the design of the study, carried
out the experiments and drafted the manuscript.
M. Jatupornpipat participated in sample analyses.
R. A. Rastall participated in the design of the study and drafting
of the manuscript.
Acknowledgements
Financial support from Ofce of the National Research Council
of Thailand, is gratefully acknowledged. We would like to thank
B. Ooraikul, Professor Emeritus, University of Alberta, for correc-
tion of the manuscript.
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