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-Glucosidase and Is Antiglycemic in Rats and Humans after Single Oral Treatments
*
Research and Development Department, Nippon Supplement, Inc., Kita-Ku, Osaka, 531-0076, Japan and
Ohshima Clinic, Ibaraki, Osaka, 567-0829, Japan
1To whom correspondence should be addressed at Research and Development Department, Nippon Supplement, Inc., 1-1-88, Oyodonaka, Kita-Ku, Osaka, 531-0076, Japan. E-mail: ngfh03{at}mail.nichigo.co.jp
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONREFERENCES |
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-glucosidase. We orally administered sucrose (2 g/kg) with or
without Touchi extract (TE) to normal rats at 100 and 500 mg/kg.
Postprandial increases in blood glucose levels at 30 and 60 min after
the administration of TE were significantly depressed compared with
controls. In humans, eight borderline diabetic subjects were
administered 0.1–10.0 g TE before sucrose loading (75 g). TE decreased
the glycemic response dose dependently after sucrose loading. Compared
with the area under the curve of the postprandial rise in blood glucose
with various doses, TE elicited a significant antiglycemic effect at a
minimum effective dose of 0.3 g. In addition, when four diabetics
were administered 0.3 g TE before eating 200 g of cooked
rice, the postprandial increases in blood glucose and mean insulin
levels were significantly depressed at 60 and 120 min, respectively,
after ingestion compared with levels when no TE was administered. TE,
which exhibits -glucosidase inhibitory activity, demonstrated an
antihyperglycemic effect and may have potential use in the management
of patients with non–insulin-dependent diabetic mellitus.
KEY WORDS: • Touchi • -glucosidase inhibitory action • antihyperglycemic effect • non–insulin-dependent diabetic mellitus • rats • humans
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONREFERENCES |
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-glucosidase inhibitory action delay carbohydrate
digestion in the small intestinal tract and thereby reduce
meal-induced rises in blood glucose and plasma insulin levels
(1
,2)
. Acarbose and voglibose are typical therapeutic
agents that display such a mechanism of action (1
2
3
4)
on
postprandial blood glucose levels and are useful oral antidiabetic
additions to the drugs currently available for the treatment of
patients with type 2 non–insulin-dependent diabetic mellitus
(3
,4)
.
The importance of biologically active substances in foods has recently
received much attention, and many physiological effects of foods
(5
6
7
8
9
10
11
12
13
14)
have been reported. The antiglycemic effects of a
variety of foods with indigestible dextrin (8)
, resistant
starch (9)
, -glucosidase (10
11
12
13)
or
-amylase inhibitory action (14)
isolated from various
sources have been investigated in animals or humans. In the present
study, we screened for -glucosidase inhibitory action in many
foodstuffs and found a water-soluble Touchi extract
(TE)2
that exhibited potent inhibitory action on rat intestinal
-glucosidase. Touchi, a traditional Chinese food, is used mainly for
seasoning. Touchi is obtained through first steaming and then
fermenting soybeans with koji (Aspergillus sp.). In addition
to the effects of TE on the postprandial rise in blood glucose after
sucrose loading in normal rats and borderline diabetic
subjects, serum insulin levels in diabetic humans were examined
in this study.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONREFERENCES |
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Touchi (100 g), obtained from commercial sources, was milled and
suspended in 900 mL of water before being boiled for 60 min. This was
followed by centrifugation at 2050 x g for 30 min at
room temperature, and the supernatant was filtered with paper (No. 5C;
Toyo Roshi, Tokyo, Japan). The filtrate was electrodialysed with
microacylizer-G3 (Asahikasei Industry Ltd., Kawasaki, Japan), and the
dialysate was concentrated before being dried under a stream of air.
The powder thus obtained was the TE used in this study. The
IC50 value of TE in rat intestinal
-glucosidase inhibition using sucrose as a substrate measured 0.34
g/L according to the method described by Miwa et al. (15)
.
In vivo studies with animals
Experiments were performed on 250- to 300-g normal male rats (Shimizu Laboratory Supplies, Kyoto, Japan). After a 12-h food deprivation, the rats were administered a sucrose solution (2 g/kg) orally with or without TE (100 and 500 mg/kg) using a stomach tube. Blood samples were collected from the tail vein and placed into heparinized (0.1 mg) tubes. Glucose levels were determined according to the oxidase method with Glucose Test Wako (Wako Pure Chemical Industries, Osaka, Japan) before (0 min) and 15, 30, 60 and 120 min after sucrose administration. Animals were treated according to National Research Council 1985 guidelines for the care and use of laboratory animals.
In vivo studies with humans
After being briefed on the experiments, volunteers gave consent for participation in the study. The protocol study was complied according to guidelines stipulated in the revised Helsinki Declaration of 1989.
Borderline diabetic subjects.
The age range of eight male borderline diabetic subjects was 29–56 y
(mean, 40.1 ± 3.0 y), and their weight was 55–80 kg (mean,
66.1 ± 2.8 kg) with a body mass index (BMI) of 19.7–28.7
kg/m2 (mean, 23.1 ± 1.2 kg/m2). Their
fasting blood glucose was <6.1 mmol/L (mean, 6.0 ± 0.2 mmol/L),
and their blood glucose level 1 h after loading with 75 g
glucose in the oral glucose tolerance test was >8.3 mmol/L (mean, 11.6
± 0.5 mmol/L). After a 12-h fast, subjects were orally loaded
with sucrose (75 g) with and without 0.1, 0.3, 1.0, 3.0 or 10 g
TE. Blood samples were taken by fingerprick at 0, 30, 60, 90 and 120
min after TE administration on d 0, 7, 14, 21, 28 and 35 after the
weekly administration of sucrose with or without TE. Blood glucose
levels were determined after oral administration by Glutest ACE (Sanwa
Kaken K.K., Nagoya, Japan). Because TE is an -glucosidase inhibitor,
sucrose instead of glucose would be more meaningful in this
experiment.
Diabetic patients.
The age range of the four diabetic patients (one man and three
women) was 50–64 y (mean, 59.3 ± 3.9 y), and their weight
was 52–74 kg (mean, 62.5 ± 4.5 kg) with a BMI of 23.4–26.2
kg/m2 (mean, 25.1 ± 0.6 kg/m2). Their
fasting blood glucose levels were >6.0 mmol/L (mean, 11.6 ± 1.6
mmol/L), and glycated hemoglobin (HbA1c) levels were >6.5% (mean 8.8
± 1.3%). After a 12-h fast, patients consumed 200 g of rice
(
1260 kJ) with or without TE (0.3 g). Blood glucose and insulin
levels were determined before (0 min) and 30, 60, 90 and 120 min after
rice consumption with or without TE administration. Rice was used in
our study because it has been used extensively in human studies of
diabetes (14
,15)
and is served as a daily staple food in
the Japanese population.
Statistical analysis
Results are expressed as means ± SEM. All data
were initially analyzed by ANOVA for each group. When a significant
F-value (P < 0.05) was obtained, this test
was followed by the Duncan’s multiple range test (SAS Institute,
Tokyo, Japan). The area under the curve was calculated using the
trapezoidal method (16)
.
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RESULTS AND DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONREFERENCES |
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At a dose of 500 mg/kg, TE significantly (P < 0.
01) decreased the postprandial rise in blood glucose compared with the
control group (n = 10) at 15–60 min after sucrose
loading (Fig. 1
). In addition, TE at a lower dose of 100 mg/kg significantly
(P < 0.05) depressed the postprandial rise in blood
glucose compared with the control group at 30 min after sucrose
loading. Furthermore, the TE-treated groups (100 and 500 mg/kg) had
significantly (P < 0.01) lower postprandial rises in
blood glucose levels (17.96 ± 0.42 and 16.76 ± 0.39 h
· mmol/L for 100 and 500 mg/kg TE, respectively) compared with the
control group (19.91 ± 0.58 h · mmol/L) when
area-under-the-curve values were compared. TE significantly depressed
the postprandial rise in blood glucose after oral sucrose loading in a
dose-dependent manner.
|
We previously used biochemical and blood analyses (data not shown) to
confirm the safety of the TE (10 g) with single oral bolus
administration in four healthy volunteers. In borderline diabetic
subjects who were administered TE, postprandial increases after oral
sucrose loading were markedly reduced (Table 1
). At doses of 
0.3 g, TE significantly suppressed the postprandial
blood glucose levels compared with the controls at 60 and 90 min after
sucrose loading. Furthermore, TE manifested dose-dependent
antiglycemic effects after sucrose administration based on evaluation
of the area-under-the-curve data (Table 1)
. The minimum effective dose
was 0.3 g.
|
). Both borderline diabetics and diabetic patients had no complaints of
any side effects such as abdominal pain, diarrhea, retching or
flatulence after TE ingestion. Moreover, abnormalities in hematological
and biochemical blood data were not observed.
|
-glucosidase using maltose as a substrate was 1.1 g/L
(data not shown). In addition, TE (100 mg/kg orally) exhibited
significant antiglycemic effects after maltose (2 g/kg) administration
(in our preliminary studies). However, TE does not show any inhibitory
activity against -amylases (data not shown) and therefore has
specific action against the enzyme -glucosidase. Because of this
enzyme-specific action, TE may have elicited such an excellent
outcome without inducing any side effects such as diarrhea, retching
and flatulence, which are commonly encountered with the use of
currently available -glucosidase inhibitory therapeutic agents.
Moreover, such favorable outcomes may be due to the lower potency of TE
in inhibition of -glucosidase in the small intestinal tract compared
with currently used -glucosidase inhibitory therapeutics
(IC50 value of voglibose, an -glucosidase
inhibitory drug, was 11 pmol/L in our assay system). However, acarbose,
an -glucosidase inhibitor, was used in long-term (1 y) studies
in diabetic patients and resulted in the passage of some dietary
carbohydrate into the colon, although a significant loss of energy into
the feces was not induced (17)
. Utilization of
carbohydrates by colonic bacteria induces the production of hydrogen
and short-chain fatty acids, which are inversely correlated with
proliferation in the rectal upper crypt, a biomarker of risk for
colonic neoplasia (17)
. TE ingestion may be used on a
long-term basis without producing abdominal distention. Through
timely regimen and proper dosing, TE may be useful for the management
of borderline diabetics and diabetic patients. Before its therapeutic
use in humans is actualized, studies on drug tolerance and the
antiglycemic effects of long-term treatment with TE are warranted.
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ACKNOWLEDGMENTS |
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-glucosidase inhibitory
assay and preliminary animal studies. |
FOOTNOTES |
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Manuscript received October 2, 2000. Revision accepted December 19, 2000.
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REFERENCES |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONREFERENCES |
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Miwa I., Okuda J., Horie T., Nakayama M. Inhibition of intestinal -glucosidases and sugar absorption by flavanes. Chem. Pharm. Bull. 1986;34:838-844
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) or with
100 mg/kg (
) or 500 mg/kg (
). Values are means ± SEM, n = 10. *P < 0.05 and **P < 0.01 versus controls.