Extraction, purification, characterization and hypoglycemic activity of a polysaccharide isolated from the root of Ophiopogon japonicus

Article abstract of DOI:10.1016/j.carbpol.2010.08.050

In this research, a water-soluble polysaccharide (OJP1) extracted with hot water from the roots of Ophiopogon japonicus which is a traditional Chinese medicinal herb was precipitated with 95% ethanol and purified by DEAE-52 cellulose anion-exchange and Sephadex G-100 gel filtration chromatography. The high performance gel permeation chromatography (HPGPC) analysis showed that the average molecular weight (Mw) of OJP1 was 35.2 kDa. Monosaccharides analysis revealed that the OJP1 is composed of Ara, Glc, Gal with a relative molar ratio of 1:16:8. Pharmaceutical experiments showed OJP1 can significantly reduce blood glucose level, increase the insulin level and remediating destruction of pancreatic islets in STZ-induced diabetic rats compared with the diabetic control group. The study shows that OJP1 has an anti-diabetic effect on STZ-induced diabetic rats, and has potential use as an anti-diabetic agent. Copyright

Full text of DOI:10.1016/j.carbpol.2010.08.050

Carbohydrate Polymers 83 (2011) 749–754
Contents lists available at ScienceDirect
Carbohydrate Polymers
journal homepage: www.elsevier.com/locate/carbpol
Extraction, purification, characterization and hypoglycemic activity of a
polysaccharide isolated from the root of Ophiopogon japonicus
Xiaoming Chena,∗,1, Jing Jina,1, Jia Tanga, Zhongfu Wangb, Junjun Wanga,
Liqin Jina, Jianxin Lua,∗
a Institute of Glycobiological Engineering/Zhejiang Provincial Key Laboratory of Medical Genetics, Wenzhou Medical College, Wenzhou 325035, China
b Key Laboratory of Ministry of Education for Western China Resource Biology and Biotechnology, College of Life Science, Northwest University, Xi’an 710069, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 2 June 2010
Received in revised form 14 August 2010
Accepted 22 August 2010
Available online 27 August 2010
In this research, a water-soluble polysaccharide (OJP1) extracted with hot water from the roots of Ophio-
pogon japonicus which is a traditional Chinese medicinal herb was precipitated with 95% ethanol and
purified by DEAE-52 cellulose anion-exchange and Sephadex G-100 gel filtration chromatography. The
high performance gel permeation chromatography (HPGPC) analysis showed that the average molecular
weight (Mw) of OJP1 was 35.2 kDa. Monosaccharides analysis revealed that the OJP1 is composed of Ara,
Glc, Gal with a relative molar ratio of 1:16:8. Pharmaceutical experiments showed OJP1 can significantly
reduce blood glucose level, increase the insulin level and remediating destruction of pancreatic islets in
STZ-induced diabetic rats compared with the diabetic control group. The study shows that OJP1 has an
anti-diabetic effect on STZ-induced diabetic rats, and has potential use as an anti-diabetic agent.
Crown Copyright © 2010 Published by Elsevier Ltd. All rights reserved.
Keywords:
Ophiopogon japonicus
Polysaccharide
Characterization
Hypoglycemic activity
1. Introduction
in traditional medicine (Kim et al., 2006; Li, Zheng, Bukuru, & De
Kimpe, 2004; Lo, Tsai, Wasser, Yang, & Huang, 2006; Kesari, Gupta,
Diabetes is a hereditary, chronic metabolic disease charac-
terized by hyperglycaemia that results from absolute or relative
deficiency of insulin secretion, impaired insulin action, or both. It
is a group of heterogeneous, autoimmune, hormonal and metabolic
disorders, often accompanied by hyperphagia (obesity), a selec-
tive loss of pancreatic islet ␤-cell mass, high blood glucose level,
and microvascular complications (Adikwu, Yoshikawa, & Takada,
2004; Yamac et al., 2008). It is the third most life threatening
disease whose mortality is right after cancer and cardiovascular
disease. Consequently, diabetes is rapidly emerging as a global
health care problem that threatens to reach pandemic levels by
2030; the number of people with diabetes worldwide is projected
to increase from 171 million in 2000 to 366 million by 2030 (Lin,
Jiang, Hu, Qiao, & Tuo, 2007; Wild, Roglic, Green, Sicree, & King,
2004). Research and development of drugs against diabetes and its
complications have been getting more and more attentions. How-
ever, currently available drugs for diabetes mellitus have a number
of limitations such as adverse effects, limited efficacy and high
rates of secondary failure. Therefore, there is a strong incentive to
develop new hypoglycemic agents, and the search for appropri-
ate hypoglycemic agents has recently focused on many plants used
& Watal, 2005).
Ophiopogon japonicus (Thunb.) Ker-Gawl, widely distributed in
South-east Asia, is a traditional Chinese medicine used to treat car-
diovascular and chronic inflammatory diseases for thousands of
years (Xiao, 2002), and has been confirmed in various experiments
as having anti-inflammatory, anti-arrhythmia, and microcircula-
tion improvement, etc. (Zhou et al., 2003; Huang & Ni, 2003).
Chemical studies have shown that this plant includes saponins,
polysaccharide and homoisoflavonoidal compounds (Kou et al.,
2005). The polysaccharides isolated from the roots of O. japoni-
cus have been reported to exhibit a variety of biological activities,
including immunostimulation, anti-ischaemia, inhibiting platelets
aggregation, hypoglycemic, etc. (Zheng, Feng, Xu, Lin, & Chen,
2009; Fan & Zhang, 2006). However, there are few reports on its
hypoglycemic properties, and the structure and function of these
polysaccharides have never been well characterized. Chen, Qian,
and Wang (1998) reported a crude O. japonicus polysaccharide
without further purification had significant antagonistic effect on
the experimental hyperglycemias in normal and alloxan-induced
diabetic mice.
In the present study, we isolated, purified and characterized
the composition of the polysaccharide (designated OJP1 below)
from O. japonicus and investigated its hypoglycemic effect on
streptozotocin-induced diabetic rats. The results showed that OJP1
(Mw = 35.2 kDa) was comprised of Ara, Glc and Gal in the ratio of
1:16:8. Pharmaceutical experiments indicated that OJP1 can signif-
∗
Corresponding authors. Tel.: +86 577 86699652; fax: +86 577 86689717.
E-mail address: xmchen01@163.com (X. Chen).
1
These authors contributed equally to this work.
0144-8617/$ – see front matter. Crown Copyright © 2010 Published by Elsevier Ltd. All rights reserved.
doi:10.1016/j.carbpol.2010.08.050

750
X. Chen et al. / Carbohydrate Polymers 83 (2011) 749–754
icantly reduce blood glucose level and increase the insulin level in
streptozotocin-induced diabetic rats, and has potential use as an
anti-diabetic agent.
2.4. General methods
Evaporation was performed at around 45 ◦C under reduced
pressure. The products were dried by lyophilization. Gas chro-
matography (GC) (Shimadzu GC-2010) equipped with RTX-50
column (30.0 m × 0.25 mm × 0.25 ␮m) and flame-ionization detec-
tor (FID). The operation was performed using the following
conditions: column temperature was programmed from 140 ◦C
(maintained for 2 min) to 170 ◦C at a rate of 6 ◦C/min, and increased
to 173 ◦C at a rate of 0.2 ◦C/min, then increased to 233 ◦C at
a rate of 6 ◦C/min, held for 40 min at 233 ◦C; the rate of N2
carrier gas was 1.0 ml/min; injection temperature was 250 ◦C;
detector temperature was 300 ◦C. Gas chromatography–mass
spectrometry (GC–MS) was run on the instrument Shimadzu
GCMS-QP2010 (Shimadzu, Japan) and equipped with RTX-50
column (30 m × 0.25 mm × 0.25 ␮m), and at temperatures pro-
grammed from 140 ◦C (maintained for 2 min) to 250 ◦C (kept for
20 min) at a rate of 3 ◦C/min. Nitrogen was the carrier gas. The
polysaccharide was monitored by the phenol–sulfuric acid method
(Dubois, Gilles, Hamilton, Rebers, & Smith, 1956).
2. Experimental
2.1. Materials and chemicals
The roots of O. japonicus were collected in Zhejiang province
(China). Sephadex G-100, DEAE-cellulose, glucose, galactose, ara-
binose, rhamnose, mannose, lactic acid, trifluoroacetic acid (TFA),
streptozotocin (STZ) were purchased from Sigma. RPMI-1640
medium (Gibco). Blood Glucose Meters was purchased from
Life scan (Milpitas, CA, USA). Insulin kits were purchased from
Changfeng Biotechnology Co. Ltd. (Zhejiang, China). Immunohis-
tochemistry kits were purchased from Zhongshan Goldenbridge
Biotechnology Co. Ltd. (Beijing, China). All the other chemicals used
were of analytical grade.
2.2. Experiment animal
2.5. Isolation and purification of the polysaccharide
Sprague–Dawley rats (body weight 250 20 g) used for exper-
iments were purchased from Experimental Center, Wenzhou
Medical College, Wenzhou, Zhejiang, China. The rats were accli-
matized for 1 week before being used for the experiment. Before
and during the experiment the rats were housed under controlled
environmental conditions of temperature (22 2 ◦C) and a 12 h
light and dark cycle, and maintained on (unless otherwise stated)
standard food pellets and tap water ad libitum.
All animal (used in this experiment) handling procedures were
performed in strict accordance with the P.R. China legislation the
use and care of laboratory animals, with the guidelines established
by Institute for Experimental Animals of Wenzhou Medical College,
and were approved by the College committee for animal experi-
ments.
The roots of O. japonicus were soaked with 95% ethanol to
remove the pigments and small lipophilic molecules. The residue
was then extracted with 10 vol. of distilled water at 90 ◦C for
3 h thrice. All water-extracts were combined, filtrated, concen-
trated, and precipitated with 95% EtOH (1:4, v/v) at 4 ◦C for
overnight. The precipitate was collected by centrifugation and
deproteinated by Sevag method (Staub, 1965). Finally the super-
natant was lyophilized to give crude polysaccharides.
The crude polysaccharides were purified by DEAE-52 cellulose
and Sephadex G-100, and the main polysaccharide fraction (OJP1)
was collected and lyophilized. OJP1 was used for further study.
2.6. Homogeneity and molecular weight
2.3. Experimental design
The homogeneity and molecular weight of OJP1 was evaluated
and determined by high performance gel permeation chromatog-
raphy (HPGPC). The sample solution was applied to Waters High
Performance Liquid Chromatography (HPLC) equipped with a
TSK-GEL G3000 SWXL column (7.8 mm × 300 mm), eluted with
0.1 mol/L Na2SO4 solution at a flow rate of 0.4 ml/min and detected
by a Waters 2414 Refractive Index Detector. The columns were cal-
ibrated with Dextran T-series standard of known molecular weight
(200,000, 70,000, 40,000, 10,000, 5000 Da). The molecular weight
of OJP1 was estimated by reference to the calibration curve made
above.
The diabetic rats were induced by the intraperitoneally (i.p.)
injection of STZ freshly at a dose of 60 mg/kg body weight. Three
days after STZ treatment, sera were collected for measurement of
blood glucose from the tail vein. The rats that were marked hyper-
glycemia (the blood glucose level > 16.7 mmol/L) were used as the
diabetic rats for further study.
The STZ-induced diabetic rats (mentioned above) were ran-
domly divided into four groups (8 rats per group), and normal rats
were used as the control.
Group I (n = 8): normal control (NC), normal rats were allowed
to free access to a normal diet and treated with saline for 28 days.
Group II (n = 8): diabetic control (DC), the diabetic rats were
allowed to free access to a normal diet and treated with saline for
28 days.
2.7. Analysis of monosaccharide composition
The monosaccharide of OJP1 was analyzed by GC. OJP1 was
hydrolyzed with 2 M TFA (2 ml) at 120 ◦C for 2 h. After removing TFA
with methanol, the hydrolyzed product was reduced with NaBH4
(50 mg), followed by neutralization with dilute acetic acid and
evaporated at 45 ◦C. The reduced products (alditols) were added
with 1 ml pyridine and 1 ml acetic anhydride in a boiling water
bath for 1 h. The acetylated products were analyzed by GC.
Group III(n = 8): OJP1-150, the diabetic rats were put on a normal
diet and treated with 150 mg/kg of OJP1 for 28 days.
Group IV (n = 8): OJP1-300, the diabetic rats were put on a nor-
mal diet and treated with 300 mg/kg of OJP1 for 28 days.
Group V (n = 8): Metformin, the diabetic rats were put on a nor-
mal diet and treated with 200 mg/kg of metformin for 28 days.
On the last day of experiment, the animals were deprived of
food overnight and sacrificed by cervical dislocation. Blood was col-
lected in polystyrene tubes without the anticoagulant. Serum was
immediately separated by centrifugation at 3000 rpm at room tem-
perature for 10 min. Samples were stored at −70 ◦C until assayed.
The pancreas were removed quickly, cleaned and washed in ice-
cold normal saline for biochemical study.
2.8. Methylation analysis
OJP1 (20 mg) was methylated three times according to the
method of Needs and Selvendran (1993). The methylated products
were extracted by chloroform and examined by IR spectroscopy.
The absence of the absorption peak corresponding to hydroxyl indi-
cated the complete methylation. The product was hydrolyzed using

X. Chen et al. / Carbohydrate Polymers 83 (2011) 749–754
751
Fig. 1. GC profile of OJP1 ((A) standard monosaccharides, peaks from left to right: Rha, Fuc, Ara, Xyl, Inositol, Man, Glc, Gal, GlcA, GalA; (B) monosaccharide composition of
OJP1, peaks from left to right: Ara, Glc, Gal).
2 M TFA, followed by reduction using NaBH4 and finally acetylated
with acetic anhydride. The partially methylated alditol acetates
were analyzed by GC–MS.
3. Results and discussion
3.1. Isolation and characterization of OJP1
The water-soluble crude polysaccharides were obtained from
the root of O. japonicus by hot water extraction, ethanol pre-
cipitation, deproteinized, and lyophilized. The extracts were
purified through with DEAE-cellulose column and Sephadex
G-100 column. The main fraction was collected and named
as OJP1 for further structure characterization and bioactiv-
ity assay. OJP1 has no absorption at 280 and 260 nm in the
UV spectrum, indicating the absence of protein and nucleic
acid.
2.9. Biochemical assays
Blood glucose and insulin levels were measured with commer-
cial kits. In brief, glycemia was assessed using a One Touch II
Meter. Insulin was determined by an enzyme-linked immunosor-
bent assay using a commercial kit, based on the direct sandwich
technique in which two monoclonal antibodies are directed against
separate antigenic determinants on the insulin molecule.
The average molecular weight of OJP1 was deter-
mined as 35.2 kDa by HPGPC. Results from phenol–sulfuric
acid assay showed that OJP1 contained 98.5% carbohy-
drate.
2.10. Pancreas histology
Tissues were fixed in 10% neutral formalin for at least 24 h
and processed for light microscopic study. Each piece of pancreas
embedded in paraffin blocs was sectioned at 5 ␮m in length before
applying on slides and stained with H&E for histopathological
examination by using light microscopy. The total operative pro-
cedures complied with the standard protocols.
OJP1 was hydrolyzed by TFA into individual monosaccharides
that were further reduced and acetylated for GC analysis. The
results showed that OJP1 is composed of Ara, Glc, Gal with a relative
molar ratio of 1:16:8 (Fig. 1).
As shown in Fig. 2, the IR spectrum of OJP1 revealed a typical
major broad stretching peak around 3422.99 cm−1 for the hydroxyl
group, and the small band at around 2934.69 cm−1 was attributed
to the C–H stretching and bending vibrations. The relatively strong
absorption peak at around 1630.66 cm−1 reflects the absorption of
the C O group that is part of glycosides (Zhu et al., 2010). The wave
number between 950 and 1200 cm−1 is often called the fingerprint
of molecules because it allows the identification of major chem-
ical groups in polysaccharides: the position and intensity of the
2.11. Statistical analysis
All results were presented as mean SD Data were analyzed by
one-way ANOVA using SPSS and Dunnett’s test. P values less than
0.05 were considered significant.

752
X. Chen et al. / Carbohydrate Polymers 83 (2011) 749–754
Fig. 3. The gas chromatogram of the products derived by methylated of the OJP1:
1: 1,4-di-O-acetyl-2,3,5-tri-O-methyl-arabinose; 2: 1,5-di-O-acetyl-2,3,4,6-tetra-
O-methyl-glucose; 3: 1,5-di-O-acetyl-2,3,4,6-tetra-O-methyl-galactose; 4: 1,3,5-
tri-O-acetyl-2,4,6-tri-O-methyl-glucose; 5: 1,2,5-tri-O-acetyl-3,4,6-tri-O-methyl-
glucose; 6: 1,4,5-tri-O-acetyl-2,3,6-tri-O-methyl-glucose; 7: 1,4,5-tri-O-acetyl-
2,3,6-tri-O-methyl-galactose; 8: 1,5,6-tri-O-acetyl-2,3,4-tri-O-methyl-glucose; 9:
1,5,6-tri-O-acetyl-2,3,4-tri-O-methyl-galactose; 10: 1,4,5,6-tetra-O-acetyl-2,3-di-
O-methyl-glucose; 11: 1,3,5,6-tetra-O-acetyl-2,4-di-O-methyl-galactose.
Fig. 2. IR spectrum of the polysaccharide of OJP1 isolated from the roots of O. japon-
icus.
Table 1
The results of methylation analysis of OJP1.
bands that are specific for each polysaccharide (Fellah, Anjukandi,
Waterland, & Williams, 2009). This region is dominated by ring
vibrations overlapped with stretching vibrations of (C–OH) side
groups and the (C–O–C) glycosidic band vibration. The absorp-
tions at 1055.31 and 1137.41 cm−1 indicated a pyranose form of
sugars (Zhao, Kan, Li, & Chen, 2005). The bands in the range of
350–600 cm−1 are assigned to skeletal modes of pyranose rings
(Yang & Zhang, 2009). On the basis of the aforementioned results, it
can be concluded that OJP1 was composed of pyranose form sugars.
The fully methylated OJP1 was hydrolyzed with acid, converted
into alditol acetates and analyzed by GC–MS (Fig. 3). The results
indicate that the polysaccharide exhibit a highly complex nature
(Table 1). The ratios of methylated fragments were calculated based
on the areas of the methylated products and corrected using the
effective-carbon response method (Sweet, Shapiro, & Albersheim,
1975). The GC–MS results (Table 1) indicated that 2,3,4-Me-Glc
(1,6-linked Glc), 2,3,6-Me-Glc (1,4-linked Glc), and 2,3-Me-Glc
(1,4,6-linked Glc) were major components of the backbone struc-
ture, part of Glc and Gal were distributed in branches, and residues
of branches terminated with Ara, Glc and Gal.
Peak No.
Methylated sugara
Molar ratio
Linkages types
1
2
3
4
5
6
7
8
9
2,3,5-Me-Ara
2,3,4,6-Me-Glc
2,3,4,6-Me-Gal
2,4,6-Me-Glc
3,4,6-Me-Glc
2,3,6-Me-Glc
2,3,6-Me-Gal
2,3,4-Me-Glc
2,3,4-Me-Gal
2,3-Me-Glc
2
5
4
4
3
5
2
7
4
7
4
Ara-(1→
Glc-(1→
Gal-(1→
→3)-Glc-(1→
→2)-Glc-(1→
→4)-Glc-(1→
→4)-Gal-(1→
→6)-Glc-(1→
→6)-Gal-(1→
→4,6)-Glc-(1→
→3,6)-Gal-(1→
10
11
2,4-Me-Gal
a
2,3,4,6-Me-Glc = 1,5-di-O-acetyl-2,3,4,6-tetra-O-methyl-glucose, etc.
atic ␤-cells by alkylation of DNA resulting in reduced synthesis and
release of insulin (Montilla et al., 2004).
The effects of OJP1 on blood glucose level in STZ-induced dia-
betic rats were shown in Table 2. The blood glucose level in normal
rats maintained constant during 4 weeks and was significantly
(P < 0.01) lower than those of STZ-induced diabetic rats of the rest
four groups. The daily administration of OJP1 (150, 300 mg/kg)
in STZ-induced diabetic rats caused a significant reduction in the
blood glucose level when compared with the diabetic control group
(P < 0.05). At fourth week, the mean decrease percentage of blood
glucose levels caused by OJP1 at the dose of 150 mg/kg was 32.9%.
3.2. Hypoglycemic activity of OJP1
Streptozotocin is one of the most commonly used substances to
induce diabetes in the rat. This toxin causes the death of pancre-
Table 2
The effect of OJP1 on blood glucose levels in STZ-induced diabetic rats.a
Blood glucose (mmol/l)
0 Days
7 Days
14 Days
21 Days
28 Days
Normal control
Diabetic control
OJP1-150
OJP1-300
Metformin
4.45 0.28
19.25 1.56b
20.03 2.07b
20.22 0.79b
19.82 1.95b
5.37 0.30
20.47 2.16b
15.72 1.98c
19.68 2.22
16.82 2.72d
4.83 0.60
23.97 2.57b
14.45 2.43c
18.58 3.79d
15.67 3.02c
5.78 0.39
21.15 2.10b
12.83 2.54c
15.92 4.35c
14.25 2.90c
5.30 0.41
22.43 2.18b
13.45 2.69c
17.32 3.83d
15.45 3.09c
a
Data represent mean SD (n = 8 for each group).
P < 0.01 (compared with the normal control).
P < 0.01 (compared with the diabetic control).
P < 0.05 (compared with the diabetic control).
b
c
d
Table 3
The effect of OJP1 on insulin levels in STZ-induced diabetic rats.a
Normal control
Diabetic control
9.79 0.62b
OJP1-150
OJP1-300
Metformin
Insulin (IU/ml)
18.24 1.22
20.79 1.42b,c
11.44 0.86b,d
17.70 1.03c
a
Data represent mean SD (n = 8 for each group).
P < 0.01 (compared with the normal control).
P < 0.01 (compared with the diabetic control).
P < 0.05 (compared with the diabetic control).
b
c
d

X. Chen et al. / Carbohydrate Polymers 83 (2011) 749–754
753
Fig. 4. Effects of OJP1 on pancreas of STZ-induced diabetic rats: (A) nondiabetic animals given saline as the negative control, (B) diabetic animals given saline as the diabetic
control, (C) diabetic animals given OJP1 at a dose of 150 mg/kg, and (D) diabetic animals given OJP1 at a dose of 300 mg/kg. Magnification ×200.
The results showed that lower dose (OJP1-150) was a little more
effective in reducing blood glucose level than Metformin on STZ-
induced diabetic rats. However, no significant differences were
observed between OJP1-150 group and Metformin group.
determined from the present study, the effects on blood glucose
and insulin and the effects on pancreatic ␤ cells could all be the
indirect result of OJP1’s action.
The effects of OJP1 on insulin level in STZ-induced diabetic rats
were shown in Table 3. The insulin level in diabetic control rats
was significantly (P < 0.01) lower than that of normal control rats.
After 4 weeks, insulin levels increased significantly in OJP1-traeted
groups and metformin-treated group (P < 0.05).
Pancreatic ␤ cells are highly specialized cells which are respon-
sible for producing all of the insulin required by an organism to
maintain glucose homeostasis. Defects in development, mainte-
nance, or expansion of ␤-cell mass can result in impaired glucose
metabolism and diabetes (Ackermann & Gannon, 2007).
As shown in Fig. 4, the results of immunohistochemical staining
of the pancreatic tissues showed that strong insulin antigen posi-
tivity was detected in the ␤-cells of the islets in healthy rats, and
islets maintained a normal rounded appearance (Fig. 4A). In con-
trast, there was weak insulin immunoreactivity in a few ␤-cells in
the islet of diabetic control rats (Fig. 4B), many of the groupings of
insulin-positive cells had lost any of the appearance of an islet struc-
ture and were seen as single cells and the islet was disorganized.
OJP1 treatment considerably increased the insulin antigenesity of
diabetic islet ␤-cells and had a normal appearance, suggesting the
possibility of ␤-cell proliferation or regeneration by OJP1 therapy
(Fig. 4C and D). The results also showed that the number of cells
increased significantly in OJP1-treated groups, and OJP1 seemed to
have the potential of remediating destruction of pancreatic islets.
Treatment of STZ-induced diabetic rats with OJP1 can signif-
icant prevented the development of diabetes. More interesting,
the hypoglycemic effect of OJP1 was not dose-dependent, and the
mechanism is not clear yet. We postulate that the 300 mg dose may
be an overdose, and further studies will be carried out. Although
the specific mechanism of hypoglycemic activity of OJP1 cannot be
4. Conclusion
This study has demonstrated that OJP1 isolated from the root
of O. japonicus is a heteropolysaccharide consisting of Ara, Glc, Gal
with a relative molar ratio of 1:16:8, and 1,6-linked Glc, 1,4-linked
Glc and 1,4,6-linked Glc were major components of the backbone
structure, part of Glc and Gal were distributed in branches, and
residues of branches terminated with Ara, Glc and Gal.
The pharmacological assays suggest that OJP1 (150, 300 mg/kg)
administered orally in STZ-induced diabetic rats can significantly
reduce blood glucose levels, increase the insulin levels, remediat-
ing destruction of pancreatic islets and the damage of pancreatic
␤-cells. It could be concluded that OJP1 may be considered as a
potential candidate for developing a new anti-diabetic agent. The
further studies on the mechanism of hypoglycemic activity are
undergoing and will be reported in due course.
Acknowledgments
This work was supported by the National Natural Science Foun-
dation of China (No. Y507096), Natural Science Foundation of
Zhejiang Province of China (No. 20872113), and Foundation of
Wenzhou Medical College (No. QTJ07020).
References
Ackermann, A. M., & Gannon, M. (2007). Molecular regulation of pancreatic ␤-
cell mass development, maintenance, and expansion. Journal of Molecular
Endocrinology, 38, 193–206.
Adikwu, M. U., Yoshikawa, Y.,
& Takada, K. (2004). Pharmacodynamic-
pharmacokinetic profiles of metformin hydrochloride from a mucoadhesive

754
X. Chen et al. / Carbohydrate Polymers 83 (2011) 749–754
formulation of a polysaccharide with antidiabetic property in streptozotocin-
induced diabetic rat models. Biomaterials, 25, 3041–3048.
Chen, W. H., Qian, H., & Wang, H. Z. (1998). Effect of polysaccharide from Ophio-
pogonis tuber on blood sugar in normal and experimental diabetic mice. Chinese
Journal of Modern Applied Pharmacy, 15(4), 21–23.
Dubois, M., Gilles, K. A., Hamilton, J. K., Rebers, P. A., & Smith, F. (1956). Colorimetric
method for determination of sugars and related substance. Analytical Chemistry,
28, 350–356.
Fan, J., & Zhang, X. (2006). Research progress in pharmacology of Ophiopogon japon-
icus polysaccharides. Chinese Archives of Traditional Chinese Medicine, 24(4),
626–627.
Montilla, P., Barcos, M., Munoz, M. C., Munoz-Castaneda, J. R., Bujalance, I., & Tunez, I.
(2004). Protective effect of Montilla–Moriles appellation red wine on oxidative
stress induced by streptozotocin in the rat. The Journal of Nutritional Biochemistry,
15, 688–693.
Needs, P. W., & Selvendran, R. R. (1993). Avoiding oxidative degradation dur-
ing sodium hydroxide/methyl iodide-mediated carbohydrate methylation in
dimethyl sulfoxide. Carbohydrate Research, 245, 1–10.
Staub, A. M. (1965). Removal of proteins from polysaccharides. Methods in Carbohy-
drate Chemistry, 5, 5–7.
Sweet, D. P., Shapiro, R. H., & Albersheim, P. (1975). Quantitative analysis by various
g.l.c. response-factor theories for partially methylated and partially ethylated
alditol acetates. Carbohydrate Research, 40(2), 217–225.
Fellah, A., Anjukandi, P., Waterland, M. R., & Williams, M. A. K. (2009). Determining
the degree of methylesterification of pectin by ATR/FT-IR: Methodology optimi-
sation and comparison with theoretical calculations. Carbohydrate Polymers, 78,
847–853.
Wild, S., Roglic, G., Green, A., Sicree, R., & King, H. (2004). Global prevalence of dia-
betes: estimates for the year 2000 and projections for 2030. Diabetes Care, 27,
1047–1053.
Huang, H. C., & Ni, Z. (2003). Effect of Ophiopogonis on the auricular microcirculation
in mice. Shanghai Laboratory Animal Science, 23, 57–58.
Xiao, P. G. (2002). Modern Chinese Materia Medica. Beijing: Chemical Industry Press.,
pp. 77–81.
Kesari, A. N., Gupta, R. K., & Watal, G. (2005). Hypoglycemic effects of Murraya
koenigii on normal and alloxan-diabetic rabbits. Journal of Ethnopharmacology,
97, 247–251.
Kim, S. W., Hwang, H. J., Cho, E. J., Oh, J. Y., Baek, Y. M., Choi, J. W., & Yun, J. W.
(2006). Time-dependent plasma protein changes in Streptozotocin-induced dia-
betic rats before and after fungal polysaccharide treatments. Journal of Proteome
Research, 5, 2966–2976.
Yamac, M., Kanbak, G., Zeytinoglu, M., Bayramoglu, G., Senturk, H., & Uyanoglu,
M. (2008). Hypoglycemic effect of Lentinus strigosus (Schwein.) Fr. Crude
exopolysaccharide in Streptozotocin-induced diabetic rats. Journal of Medicinal
Food, 11(3), 513–517.
Yang, L., & Zhang, L. M. (2009). Chemical structural and chain conformational char-
acterization of some bioactive polysaccharides isolated from natural sources.
Carbohydrate Polymers, 76, 349–361.
Kou, J., Sun, Y., Lin, Y., Cheng, Z., Zheng, W., Yu, B., & Xu, Q. (2005). Anti-inflammatory
activities of aqueous extract from Radix Ophiopogon japonicus and its two con-
stituents. Biological & Pharmaceutical Bulletin, 28(7), 1234–1238.
Li, W. L., Zheng, H. C., Bukuru, J., & De Kimpe, N. (2004). Natural medicines used in
the traditional Chinese medical system for therapy of diabetes mellitus. Journal
of Ethnopharmacology, 92, 1–21.
Zhao, G. H., Kan, J. Q., Li, Z. X., & Chen, Z. D. (2005). Structural features and
immunological activity of a polysaccharide from Dioscorea opposita Thunb roots.
Carbohydrate Polymers, 61, 125–131.
Zheng, Q., Feng, Yi., Xu, D. S., Lin, X., & Chen, Y. Z. (2009). Influence of sulfation on anti-
myocardial ischemic activity of Ophiopogon japonicus polysaccharide. Journal of
Asian Natural Products Research, 11(4), 306–321.
Lin, G. P., Jiang, T., Hu, X. B., Qiao, X. H., & Tuo, Q. H. (2007). Effect of Siraitia grosvenorii
polysaccharide on glucose and lipid of diabetic rabbits induced by feeding high
fat/high sucrose chow. Experimental Diabetes Research, 1–4.
Lo, H. C., Tsai, F. A., Wasser, S. P., Yang, J. G., & Huang, B. M. (2006). Effects of
ingested fruiting bodies, submerged culture biomass, and acidic polysaccharide
glucuronoxylomannan of Tremella mesenterica Retz.:Fr. on glycemic responses
in normal and diabetic rats. Life Sciences, 78, 1957–1966.
Zhou, Y. H., Xu, D. S., Feng, Y., Fang, J. N., Xia, H. L., & Liu, J. (2003). Effects on nutrition
blood flow of cardiac muscle in mice by different extracts in radix Ophiopogonis.
China Journal of Experimental Traditional Medical Formulae, 9, 22–23.
Zhu, M. Y., Mo, J. G., He, C. S., Xie, H. P., Ma, N., & Wang, C. J. (2010). Extraction,
characterization of polysaccharides from lycium barbarum and its effect on bone
gene expression in rats. Carbohydrate Polymers, 80, 672–676.