Food Chemistry 130 (2012) 261–266

Contents lists available at ScienceDirect

Food Chemistry
journal homepage: www.elsevier.com/locate/foodchem

Inhibitory potential of trilobatin from Lithocarpus polystachyus Rehd

against a-glucosidase and a-amylase linked to type 2 diabetes
Hua-Qiang Dong ⇑, Mei Li, Feng Zhu, Fu-Lai Liu, Jian-Bo Huang
Department of Food Science, Foshan University, Foshan 528231, China

a r t i c l e i n f o a b s t r a c t

Article history: The chemical structure of the sweet compound from Lithocarpus polystachyus Rehd was identified as trilob-
Received 18 September 2009 atin on the basis of HPLC, EIS-MS and NMR analyses. The inhibitory activities of trilobatin against a-gluco-
Received in revised form 17 June 2011 sidase and a-amylase were evaluated, and the inhibition mechanism was analysed with Lineweaver–Burk
Accepted 8 July 2011
plots. Also the antioxidant activity evaluation of trilobatin was conducted by DPPH radical scavenging
Available online 5 August 2011
assay. Comparing with acarbose, trilobatin showed a strong inhibitory activity against a-glucosidase and
a moderate inhibitory activity against a-amylase. The Lineweaver–Burk plots analysis elucidated that tril-
Keywords:
obatin inhibited the enzyme non-competitively. DPPH scavenging activity of trilobatin (IC50 = 0.57 mg/ml)
Lithocarpus polystachyus Rehd
Trilobatin
was higher than rutin (IC50 = 0.72 mg/ml), which indicated that trilobatin had a moderate antioxidant
a-Glucosidase potential. These results suggest that trilobatin is a potential effective a-glucosidase inhibitor for manage-
a-Amylase ment of postprandial hyperglycemia with less side effect, and provide strong rationale for further animal
Type 2 diabetes and clinical studies.
Ó 2011 Elsevier Ltd. All rights reserved.

1. Introduction blood glucose levels, of type 2 diabetes (Melo, Gomes, & Carvalho,
2006). These chemical drugs have strong inhibitory activities
Diabetes is increasing rapidly in every part of the world, the prev- against both a-glucosidase and a-amylase, but have a side effects
alence of the disease will grow to 380 million people affected by such as abdominal distention, flatulence, meteorism and possibly
2025 (Silink, 2009). Of the diabetics, about 90% have type 2 diabetes diarrhoea (Bischoff, Puls, Krause, Schutt, & Thomas, 1985). Previous
(NIDDM; non-insulin-dependent diabetes mellitus) (American Dia- researches indicated that these side effects could be caused by
betes Association, 2005). Type 2 diabetes is characterised by abso- excessive inhibition of pancreatic a-amylase resulting in the
lute or relative deficiencies in insulin secretion and/or insulin abnormal bacterial fermentation of undigested carbohydrates in
action associated with chronic hyperglycemia and disturbances of the colon (Horii et al., 1987). Therefore, more effective inhibitors
carbohydrate, lipid, and protein metabolism (WHO, 1999). The for the two enzymes should have strong inhibitory effect against
chronic hyperglycemia in type 2 diabetes can be expressed in two a-glucosidase and mild inhibitory effect against a-amylase, which
indexes, fasting and postprandial blood glucose levels. Researches can be an effective therapy for managing postprandial hyperglyce-
in recent years have indicated that postprandial high blood glucose mia with minimal side effects (Kwon, Vattem, & Shetty, 2006).
level is a more important factor to result in onset and development The chronic hyperglycemia in diabetes causes oxidative stress
of type 2 diabetes (Chang, Smith, & Bloem, 2004). (Brownlee, 2005). It is believed that oxidative stress plays an impor-
One important factor to result in a postprandial hyperglycemia is tant role in chronic complications of diabetes (Dicarli, Janises, Grun-
the fast uptake of glucose in the intestine, in which a-amylase and berger, & Ager, 2003). Therefore, alleviation of oxidative stress is
a-glucosidase play important roles due to hydrolysis of starch and essential for preventing or reversing diabetic complications (DeF-
oligosaccharide (Gray, 1995). It is believed that inhibition of these ronzo, 1999), and a compound with antioxidant activity combined
enzymes can effectively control the postprandial elevation of blood with inhibitory activities against a-glucosidase and a-amylase
glucose level. Therefore, an important strategy for managing post- should be a more effective anti-diabetic agent.
prandial hyperglycemia is to inhibit a-amylase and a-glucosidase Lithocarpus polystachyus Rehd is a shrub distributed widely in
activities (Krentz & Bailey, 2005). the mountainous area in southern China. Its tender leaves have
a-Glucosidase inhibitors, such as acarbose, have been used been a traditional Chinese herb and been taken as a sweet tonic
clinically to control blood glucose levels, especially postprandial drink for several hundreds of years (China Pharmacopoeia Com-
mittee, 1999). In the course of seeking the sweet components in
the leaves, the sweet compounds were identified as flavonoids,
⇑ Corresponding author. Address: Department of Food Science, Foshan University,
and their content in the leaves was up to around 7% (Li et al.,
Dali, Nanhai, Foshan 528231, China. Tel.: +86 757 85505064.
2001; Su, Chen, Jiao, Huang, & Li, 2000). In our previous researches,
E-mail address: [email protected] (H.-Q. Dong).

0308-8146/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.foodchem.2011.07.030
262 H.-Q. Dong et al. / Food Chemistry 130 (2012) 261–266

the flavonoids were preparatively extracted and purified from L. was incubated in 96 well plates at 37 °C for 10 min. After pre-incu-
polystachyus Rehd (Dong, Ning, Cui, et al., 2007; Dong, Ning, Yu, bation, 50 ll of 5 mM p-nitrophenyl-a-D-glucopyranoside (PNPG)
et al., 2007), and the significant anti-diabetes effects of the crude solution in 0.1 M phosphate buffer (pH 6.8) was added to each well
extract were proven with alloxan induced diabetic mice (Dong, and incubated at 37 °C for another 20 min. Then the reaction was
Ning, Li, Yu, & Lin, 2006). stopped by adding 160 ll of 0.2 M NaCO3 into each well, and absor-
In this study, the chemical structure of the preparatively sepa- bance readings (A) were recorded at 405 nm by micro-plate reader
rated flavonoid was identified, and its inhibitive potential against (SpectraMaxÒ, M2/M2e, Molecular device Co., Sunnyvale, CA, USA)
a-glucosidase and a-amylase activities were studied, and the anti- and compared to a control which had 60 ll of buffer solution in place
oxidant activity was evaluated. These will provide useful data and of the extract. The a-glucosidase inhibitory activity was expressed
information for the further researches and developments for more as inhibition % and was calculated as follows:
effective anti-diabetes agents.

Inhibition ð%Þ ¼ ðAcontrol  Asample Þ=Acontrol  100

2. Materials and methods

2.1. Materials The concentration of inhibitors required for inhibiting 50% of

the a-glucosidase activity under the assay conditions was defined
Phloridzin (99.7%), 2, 20 -diphenyl-1-picrylhydrazyl (DPPH), p-nit as the IC50 value.
rophenyl-a-D-glucopyranoside (PNPG), a-glucosidase (EC 3.2.1.20), For kinetic analyses, the enzyme and the inhibitor were incu-
and porcine pancreatic amylase (EC 3.2.1.1, type VI), were pur- bated with increasing concentrations of PNPG solution. Inhibitory
chased from Sigma–Aldrich chemical company (St. Louis, USA); kinetics was analysed by Lineweaver–Burk plots.
acarbose was Bayer’s product. All the reagents used were of analyt-
ical grade, and glass-distilled water was used for the preparation of
2.4.2. a-Amylase inhibition assay
the reagents.
The a-amylase inhibitory activity was measured according to
methods described by Kim, Jeong, Wang, Lee, and Rhee (2005)
2.2. Preparation of the flavonoid from L. polystachyus Rehd with some modifications. A 40 ll of sample solution (0.2 mg of
sample or acarbose in 20 mM sodium phosphate buffer, pH 6.9
L. polystachyus Rehd leaves were harvested in the east part of with 0.006 M sodium chloride) was premixed with 200 ll of a-
Guizhou province, China, and identified by the botanists from amylase solution (1.0 U/ml in the pH 6.9 buffer), and incubated
South China Institute of Botany, the Chinese Academy of Science. at 25 °C for 10 min. After pre-incubation, 400 ll of a 0.25% starch
The crude extract of flavonoids and the purified flavonoid were solution in the pH 6.9 buffer was added to each tube to start the
made as described by Dong, Ning, Cui, et al., 2007; Dong, Ning, reaction. The reaction was carried out at 37 °C for 5 min and ter-
Yu, et al., 2007. In the whole process of separation and purification, minated by addition of 1.0 ml of the DNS reagent (1% 3,5-dinitro-
only water and ethanol were used as solvent and eluent. salicylic acid and 12% sodium potassium tartrate in 0.4 M NaOH).
The test tubes were then incubated in a boiling water bath for
2.3. Assays of identification of the separated flavonoid compound 5 min and cooled to room temperature. The reaction mixture
was then diluted after adding 10 ml distilled water and absor-
2.3.1. HPLC analysis bance was measured at 540 nm using a UV–Vis spectrophotome-
All chromatographic analyses were conducted on a Summit ter (mini 1240, Shimadzu, Japan). The control had 200 ll of buffer
HPLC system (Dionex, Germany), equipped with a C-18 reversed- solution in place of the a-amylase solution. The % inhibition was
phase column (Diamond, 150  4.6 mm i.d., 5 lm). The mobile calculated by:
phase consisted of a mixture of methanol–water (60:40, v/v), at a
flow rate of 1.5 mL/min, and the column effluent was monitored Inhibition ð%Þ ¼ ðAcontrol  Asample Þ=Acontrol  100
at 283 nm with a UV diode-array detector. The column was main-
tained at ambient temperature (25–30 °C). 2.5. DPPH radical scavenging assay

2.3.2. EIS-MS analysis Free radical scavenging activity of extracts was determined
Mass spectrometry experiments were performed on a ZQ2000 using the DPPH method described by Chen, Wu, Shieh, Kuo, and
mass spectrometer (Waters, USA), equipped with an electrospray Hsieh (2006), and Ozsoy, Can, Yanardag, and Akev (2008) with
source, operating at ESI (ve mode). MS parameters were as follows: some modifications. A 50 ll of methanol solution containing differ-
negative ionisation mode, capillary, 2.89 kV; source temperature, ent concentrations of sample or 0.2 mg/ml of positive control vita-
100 °C; desolvation temperature, 250 °C; mass range, 50–800 m/z. min C was mixed with 150 ll of freshly prepared DPPH methanol
solution (0.2 mM) in one well of a 96-well microplate. An equal
2.3.3. NMR analysis amount of methanol was used as a control. The mixture was sha-
Chemical structure of the purified compound was analysed in ken vigorously and allowed to stand in the dark at room tempera-
dimethylsulphoxide (DMSO)-d6 on a Bruker DRX-400 MHz spectro ture for 30 min. The decrease in absorbance (A) of the resulting
meter. solution was then measured at 517 nm against methanol on a mi-
cro-plate reader (SpectraMaxÒ, M2/M2e, Molecular device Co., Sun-
2.4. Assays of enzymes activity inhibition determination nyvale, CA). Activity of scavenging (%) was calculated using the
following formula:
2.4.1. a-Glucosidase assay
DPPH radical scavenging ð%Þ ¼ ðAcontrol  Asample Þ=Acontrol  100
The a-glucosidase inhibitory activity was measured as described
by Kwon, Apostolidis, and Shetty (2008), and Andrade-Cetto, Becer-
ra-Jimenez, and Cardenas-Vazquez (2008) with slight modifications. The concentration of a sample required to decrease the absor-
Briefly, a volume of 60 ll of sample solution and 50 ll of 0.1 M phos- bance at 517 nm by 50% compared to the control response was de-
phate buffer (pH 6.8) containing a-glucosidase solution (0.2 U/ml) fined as IC50 values.
 H.-Q. Dong et al. / Food Chemistry 130 (2012) 261–266 263

2.6. Statistical analysis data of the [MH] ions of both the two compounds displayed the
same features. These results indicated that the target compound
Analysis at every time point from each experiment was carried had the same chemical formula and moieties as the phloridzin stan-
out in triplicates. The results were statistically analysed by ANOVA dard, and they might be the same molecules (the glycosidic moiety
and Duncan’s multiple range tests. Statistical significance was ac- linked on the 20 -OH of the phloretin, Fig. 2A), or the isomers (the gly-
cepted at a level of p < 0.05. cosidic moiety was linked on the 40 -OH of the phloretin, Fig. 2B).

3. Results and analysis

3.1.3. NMR
The 1H and 13C NMR signals of the target compound are listed in
3.1. Chemical structure determination of the separated flavonoid
Table 1. The 1H NMR signals of d H-30 and H-50 had the same value,
compound
6.05, and the 13C NMR signals of C-30 and C-50 had the same value,
94.9, which presented that the 1H and 13C NMR signals on the two
3.1.1. HPLC
sides of C-40 were symmetric. These results identified that the gly-
The HPLC analyses of the target compound and a mixture of
cosidic moiety of the target compound was linked on the 40 -OH of
equal amounts of the target compound and the phloridzin standard
the phloretin, not on the 20 -OH, which was in concordance with the
were conducted respectively. HPLC profile of the target compound
reported results (Nie, Tanaka, Zhou, & Tanaka, 1982). Therefore, the
showed 99.87% purity and 6.28 min of retention time. In the HPLC
target compound was identified as phloretin-40 -b-D-glucoside, or
chromatography of a mixture of equal amounts of the two com-
trilobatin (Fig. 2B).
pounds, only one main peak with a retention time of 6.28 min was
observed. These results implied that the target compound had a
similar chemical structure with the phloridzin standard by HPLC. 3.2. Inhibitory effect of trilobatin (Trb) against a-glucosidase and
a-amylase
3.1.2. ESI-MS
The ESI-MS spectra of the target compound are shown in Fig. 1. As shown in Table 2, no significant differences of a-glucosidase
The molecular mass of the compound is 472 Da, and the expected inhibition rates between trilobatin (48.7%) and acarbose (45.8%),
[MH] negative ion was observed in the spectra at m/z 471. Loss were found but which were significantly higher than that of rutin
of two H2O molecules (36 Da) from this ion gave a [M2H2OH] (25.2%). On the other hand, a-amylase inhibition rate of trilobatin
ion at m/z 435. Cleavage of a glucose moiety without a OH substitu- (21.2%) was significant lower than acarbose (42.6%), and close to
ent (163 Da) from the pseudomolecular ion at 435 m/z and rutin (23.1%). These results demonstrated that Trb had the same
protonation resulted in an ion with m/z 273. These characteristic strong inhibitory activity against a-glucosidase as acarbose, and
ions can be observed in the case of the phloridzin standard, as the obviously lower inhibitory activity against a-amylase.

Fig. 1. ESI-MS spectrum of the ion [MH]of the target compound.

A B
3
5' 3 2 OH
HO 4' OH 2 OH HO
6' 5'
3' 5 O O 4' OH 5
OH 6' ß 6
2' 6
HO 3' a
O O OH
OH 2'
O
OH OH O
OH
OH

Fig. 2. Chemical structures of phloridzin (A) and trilobatin (phloretin-40 -b-D-glucoside) (B).
264 H.-Q. Dong et al. / Food Chemistry 130 (2012) 261–266

Table 1 60 Acb Trb Rutin

1
H and 13C NMR signals of the target compound (in DMSO-d6). A
Position dH dC 50

Alpha-glucosidase inhibition
1 131.4
2 7.01 129.1 40
3 6.65 115.0
4 155.4

(%)
5 6.65 115.0 30
6 7.01 129.1
C@O 205.0 20
a 45.6
b 2.77 29.3
10 105.3 10
20 163.9
30 6.05 94.9 0
40 163.3 0 0.4 0.8 1.2 1.6 2
50 6.05 94.9 Inhibitor concentrations(mg/mL)
60 163.9
G1 99.5
G2 73.0 100 Rutin Trb Vc
G3 77.1
B
G4 69.2

DPPH scavenging activity

80
G5 76.4
G6 60.4
60

(%)
40

Table 2
Inhibitory effects of trilobatin on a-glucosidase and a-amylase.a 20

Inhibitors Inhibition of a-glucosidase (%) Inhibition of a-amylase (%)

0
Acb 45.8 ± 3.4a 42.6 ± 2.5a 0 0.4 0.8 1.2 1.6 2
Trb 48.7 ± 4.3a 21.2 ± 3.2b Inhibitor concentration(mg/mL)
Rutin 25.2 ± 4.4b 23.1 ± 3.5b
a Fig. 3. a-Glucosidase inhibition (A) and DPPH radical scavenging activity (B) of
Values are the means ± SD of three replicated samples. Duncan’s multiple range
trilobatin at different concentrations. Values are the means ± SD of three replicated
test was conducted and data in the same column with different letters indicate
samples.
statistically significant differences among groups at p < 0.05.

80
3.3. Inhibitory activity against a-glucosidase with increasing [I]
concentrations of Trb
70 1.6mg
a-Glucosidase inhibitory activity was measured at 0.2, 0.4, 0.8,
1.2, 1.6, and 2.0 mg/ml of Trb, acarbose, and rutin respectively, 60 0.4mg
1/v(µM/min)

and the result is showed as Fig. 3(A). a-Glucosidase inhibitory activ-

ity increased with the concentration enhancement of the three 50
0.2mg
inhibitors from 0.2 to 0.8 mg/ml, which illustrated a good dose
dependent response. Trb had reached the highest a-glucosidase 40
Control
inhibitory activity at 0.8 mg/ml, and had even higher inhibitory
activity than acarbose at this concentration. However, based on 30
IC50 values, a-glucosidase inhibitory activity of Trb (0.37 mg/ml)
was lower than that of acarbose (0.28 mg/ml), and much higher 20
than that of rutin (0.45 mg/ml).
10

3.4. Mechanism of a-glucosidase inhibition of Trb 0

-6 -5 -4 -3 -2 -1 0 1 2 3 4 5 6 7 8 9 10 11 12 13
Fig. 4 shows a Lineweaver–Burk plot of a-glucosidase inhibitory -10 1/[pNPG](mM)
activities in presence of Trb at 0.2, 0.4 and 1.6 mg/ml and absence
of the inhibitor with different substrate concentrations of pNPG. Fig. 4. Lineweaver–Burk plot analysis of the inhibition kinetics of a-glucosidase
The linear regression and extrapolation of data gave a series of inhibitory effects by trilobatin (I).

lines crossing on the horizontal axis and the vertical axis. With
increasing trilobatin concentration, the vertical axis intercept (1/ 3.5. Antioxidant activity of Trb by DPPH radical scavenging assay
V m) increased, however, the horizontal axis intercept (1/km) re-
mained the same. These results indicate that the velocity of the The DPPH scavenging-linked antioxidant activity of Trb was
reaction catalysed by a-glucosidase was slowed down with measured while comparing with Vitamin C and rutin at concentra-
increasing trilobatin concentration, and the km value of a-glucosi- tions ranging from 0.1 to 2.0 mg/ml respectively. As shown in
dase was not affected, which was correlated with the classical pat- Fig. 3B, with concentrations increasing from 0.1 to 0.6, 1.2, and 1.6,
tern of non-competitive inhibition, and indicated that Trb and the DPPH scavenging activities of Vc, Trb and rutin increased and
substrate did not bind to a-glucosidase at the same site. reached their highest level at 88.8%, 84.2% and 81.8%, respectively.
 H.-Q. Dong et al. / Food Chemistry 130 (2012) 261–266 265

However, in the IC50 values concept, significant variance of their 5. Conclusion

DPPH scavenging activities was observed. DPPH scavenging activity
of Trb (IC50 = 0.57 mg/ml) was lower than Vc (IC50 = 0.24 mg/ml), The results from this in vitro study clearly indicated that trilob-
but higher than rutin (IC50 = 0.72 mg/ml), which indicated that Trb atin from L. polystachyus Rehd had strong inhibitory activity against
had a good in vitro antioxidant activity. a-glucosidase and mild inhibitory activity against a-amylase. The
compound had a moderate antioxidant activity, and was a strong
sweetener. These functionalities when combined in one compound
4. Discussion are potentially useful to manage the glucose-induced hyperglyce-
mia and provide the biochemical rationale for further animal and
Early effective control of postprandial hyperglycemia is impor- clinical studies.
tant in early intervention and prevention of diabetic complica-
tions for type 2 diabetes management (Ratner, 2001). a- Acknowledgements
Glucosidase and a-amylase are two key enzymes related to carbo-
hydrate digestion and elevation of levels of fasting blood glucose. The financial support provided by Guangdong Provincial Natu-
It is now believed that inhibition of these enzymes can be an ral Science Foundation (Grant No. 8152800001000023) was greatly
important strategy in the management of hyperglycemia by appreciated.
retarding the postprandial increase of blood glucose level after a
mixed carbohydrate diet (Puls, Keup, Krause, Thomas, & Hoffmei- References
ster, 1977). Currently used inhibitors of a-glucosidase and a-amy-
lase such as acarbose have strong inhibitory activities against both American Diabetes Association (2005). Diagnosis and classification of diabetes
of the two enzymes. These drug therapies are effective but have mellitus. Diabetes Care, 28, S37–S42.
side effects caused by the abnormal bacterial fermentation of Andrade-Cetto, A., Becerra-Jimenez, J., & Cardenas-Vazquez, R. (2008). Alfa-
glucosidase-inhibiting activity of some Mexican plants used in the treatment
undigested carbohydrates due to an excessive inhibition of a- of type 2 diabetes. Journal of Ethnopharmacology, 116, 27–32.
amylase. Bhandari, M. R., Jong-Anurakkun, N., Hong, G., & Kawabata, J. (2008). a-Glucosidase
Therefore, strong inhibitors of a-glucosidase with mild inhibi- and a-amylase inhibitory activities of Nepalese medicinal herb Pakhanbhed
(Bergenia ciliata, Haw). Food Chemistry, 106, 247–252.
tory activity against a-amylase have long been sought to overcome Bischoff, H., Puls, W., Krause, H. P., Schutt, H., & Thomas, G. (1985). Pharmacological
this challenge. It was reported (Kwon et al., 2008) that phenolicen- properties of the novel glucosidase inhibitors BAY m1099 (miglitol) and BAY o
riched extracts from several eggplant samples (e.g., White pulp and 1248. Diabetes Research and Clinical Practice, 1, 53–62.
Brownlee, M. (2005). The pathobiology of diabetic complications. Diabetes, 54,
Graffiti skin) have high a-glucosidase inhibitory activity combined
1615–1625.
with low two times a-amylase inhibitory activity. Bhandari, Jong- Chang, A. M., Smith, M. J., & Bloem, C. J. (2004). Effect of lowing postprandial
Anurakkun, Hong, and Kawabata (2008) and Shobana, Sreerama, hyperglycemia on insulin secretion in older people with impaired glucose
and Malleshi (2009) evaluated the inhibitory activities against a- tolerance. American Journal of Physiology–Endocrinology and Metabolism, 287,
E906–E910.
glucosidase and a-amylase of phenolic compounds extracted from Chen, F., Wu, A., Shieh, P., Kuo, D., & Hsieh, C. (2006). Evaluation of the antioxidant
several plants, they found that the phenolics inhibited both en- activity of Ruellia tuberosa. Food Chemistry, 94, 14–18.
zymes significantly with low IC50 values for a-glucosidase, but China Pharmacopoeia Committee (1999). Pharmacopoeia of the People’s Republic of
China (first part of the 2000 ed.) (pp. 327–329). Beijing: China Chemical Industry
with much higher IC50 values a-amylase. Our results indicated that Press.
flavonoid trilobatin from L. polystachyus Rehd showed significant Dicarli, M. F., Janises, J., Grunberger, J., & Ager, J. (2003). Role of chronic
inhibitory activities against a-glucosidase and a-amylase. How- hyperglycemia in the pathogenesis of coronary microvascular dysfunction in
diabetes. Journal of the American College of Cardiology, 41, 1387–1393.
ever the inhibition rate for a-glucosidase was close to that of ascar- DeFronzo, R. A. (1999). Pharmacologic therapy for type 2 diabetes mellitus. Annals of
bose, and the inhibition rate for a-amylase was obviously lower Internal Medicine, 131, 281–303.
than that of acarbose (Table 2 and Fig. 3A). This indicated that Dong, H., Ning, Z., Li, L., Yu, L., & Lin, L. (2006). Anti-hyperglycemia and effects of
flavonoid phloridzin from Lithocarpus polystachyus Rehd on diabetic model
trilobatin was a strong inhibitor for a-glucosidase with mild inhib- mice. Food Science, 12, 714–717.
itory activity against a-amylase. Dong, H., Ning, Z., Cui, Z., Li, L., Yu, L., & Lin, L. (2007). Microwave-assisted ext
On the other hand, oxidative stresses play important roles in raction of flavonoids from tender leaves of Lithocarpus polystachyus Rehd.
Transactions of the CSAE, 23(2), 213–217.
initiating beta-cell damage and insulin resistance. Anti-oxidants
Dong, H., Ning, Z., Yu, L., Li, L., Lin, L., & Huang, J. (2007). Preparative separation and
may prevent the progressive impairment of pancreatic beta-cell identification of flavonoid phloridzin from the crude extract of Sweet Tea
function and thus reduce the occurrence of type 2 diabetes (Song, (Lithocarpus polystachyus Rehd). Molecules, 12, 552–562.
Manson, Buring, Sesso, & Liu, 2005). Researchers gave much more Gray, D. M. (1995). Carbohydrate digestion and absorption—role of small intestine.
New England Journal of Medicine, 29, 1225–1230.
attention to antioxidant activities when antidiabetic activities of Horii, S., Fukasse, K., Matsua, T., Kameda, K., Asano, N., & Masui, Y. (1987). Synthesis
phytochemicals, especially phenolics, were evaluated, and it was and a-D-glucosidase inhibitory activity of N-substituted valiolamine
reported that several phenolic compounds with inhibitory activi- derivatives as potent oral antidiabetic agents. Journal of Medicinal Chemistry,
29, 1038–1046.
ties against a-glucosidase and a-amylase had moderate antioxi- Kim, Y., Jeong, Y., Wang, M., Lee, W., & Rhee, H. (2005). Inhibitory effect of pine
dant activity (Bhandari et al., 2008; Kwon et al., 2008; Shobana extract on a-glucosidase activity and postprandial hyperglycemia. Nutrition, 21,
et al., 2009). Trilobatin showed a distinct DPPH radical scavenging 756–761.
Kinghorn, A. D. (1987). Biologically active compounds from plants with reputed
activity in our results (Fig. 3B). medicinal and sweetening properties. Journal of Natural Products, 50(6),
Further, L. polystachyus Rehd is a traditional Chinese herb and 1009–1024.
its Leaves are also called Sweet Tea, which have been taken as a Krentz, A. J., & Bailey, C. J. (2005). Oral antidiabetic agents: Current role in type 2
diabetes mellitus. Drugs, 65, 385–411.
folk sweet tea in southern China for hundreds of years without Kwon, Y.-I., Vattem, D. V., & Shetty, K. (2006). Evaluation of clonal herbs of
evidence of adverse effects or toxicity. Nie et al. (1982) conducted Lamiaceae species for management of diabetes and hypertension. Asian Pacific
the isolation of sweet principles of leaves of Lithocarpus litseifolius Journal of Clinical Nutrition, 15, 107–118.
Kwon, Y. I., Apostolidis, E., & Shetty, K. (2008). In vitro studies of eggplant (Solanum
(Hance) Rehd and identified chemical structure of trilobatin.
melongena) phenolics as inhibitors of key enzymes relevant for type 2 diabetes
Kinghorn (1987) reported that the trilobatin was 300 times sweet- and hypertension. Bioresource Technology, 99, 2981–2988.
er than sugar. We preparatively separated trilobatin and identified Li, W., Li, R., Nai, Z., Na, X., Su, X., & Chen, Z. (2001). Studies on active anti-allergic
its chemical structure, thus trilobatin from L. polystachyus Rehd can constituents in Lithocarpus polystachyus (Wall) Rehd. Journal of Yunnan
University, 23, 461–463.
be a potential natural sweetener with health functions and without Melo, E. B., Gomes, A. S., & Carvalho, I. (2006). a- and b-glucosidase inhibitors:
energy for the human body. Chemical structure and biological activity. Tetrahedron, 62, 10277–10302.
266 H.-Q. Dong et al. / Food Chemistry 130 (2012) 261–266

Nie, R., Tanaka, T., Zhou, J., & Tanaka, O. (1982). Phlorizin and trilobatin, sweet Silink, M. http://www.eatlas.idf.org/newsc269.html, accessed August 2009.
dihydrochalcone-glucosides from Lithocarpus litseifolius (Hance) Rehd. Song, Y., Manson, J. E., Buring, J. E., Sesso, H. D., & Liu, S. (2005). Association of
(Fagaceae). Agricultural and Biological Chemistry, 46(7), 1933–1934. dietary flavonoids with risk of type 2 diabetes, and markers of insulin resistance
Ozsoy, N., Can, A., Yanardag, R., & Akev, N. (2008). Antioxidant activity of Smilax and systemic inflammation in women: A prospective study and cross sectional
excelsa L. Leaf extracts. Food Chemistry, 110, 571–583. analysis. Journal of the American College of Nutrition, 24(5), 376–384.
Puls, W., Keup, U., Krause, H., Thomas, P. G., & Hoffmeister, F. (1977). Glucosidase Su, X., Chen, Z., Jiao, B., Huang, Q., & Li, W. (2000). Isolation and identification of
inhibition: A new approach to the treatment of diabetes, obesity and active anti-allergic constituents in Lithocarpus polystachyus Rehd. Journal of
hyperlipoproteinemia. Naturwissenschaften, 64, 536–537. Southwest Agricultural University, 22, 419–420.
Ratner, R. E. (2001). Controlling postprandial hyperglycemia. American Journal of Word Health Organization (1999). Definition, diagnosis and classification of
Cardiology, 88, 26H–31H. diabetes mellitus and its complications. Report of WHO consultation. Geneva,
Shobana, S., Sreerama, Y. N., & Malleshi, N. G. (2009). Composition and enzyme p. 66.
inhibitory properties of finger millet (Eleusine coracana L.) seed coat phenolics:
Mode of inhibition of a-glucosidase and pancreatic amylase. Food Chemistry.
doi:10.1016/j.foodchem.2009.01.042.