Synthesis and inhibition of α-glucosidase of methyl glycyrrhetinate glycosides

Article abstract of DOI:10.1080/14786419.2019.1639181

The synthesis of the methyl glycyrrhetinate glycosides and inhibition of α-glucosidase were studied. The carboxyl group of glycyrrhetinic acid was methylated, and glucose and galactose were introduced into the hydroxyl group to obtain compounds 7 and 12.

Full text of DOI:10.1080/14786419.2019.1639181

Natural Product Research
Formerly Natural Product Letters
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Synthesis and inhibition of α-glucosidase of methyl
glycyrrhetinate glycosides
Wei Zhang, He-Ying Wang, Huai-Xu Wang & Zhen-Yuan Zhu
To cite this article: Wei Zhang, He-Ying Wang, Huai-Xu Wang & Zhen-Yuan Zhu (2019):
Synthesis and inhibition of α-glucosidase of methyl glycyrrhetinate glycosides, Natural Product
Research, DOI: 10.1080/14786419.2019.1639181
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NATURAL PRODUCT RESEARCH
https://doi.org/10.1080/14786419.2019.1639181
Synthesis and inhibition of a-glucosidase of methyl
glycyrrhetinate glycosides
a,b,c
a,b,c
Wei Zhanga,b,c , He-Ying Wang
, Huai-Xu Wang
and
ꢀ
ꢀ
Zhen-Yuan Zhua,b,c
aState Key Laboratory of Food Nutrition and Safety, Tianjin University of Science & Technology,
b
Tianjin, P.R. China; Key Laboratory of Food Nutrition and Safety, Ministry of Education, Tianjin
c
University of Science and Technology, Tianjin, P.R. China; College of Food Science and
Biotechnology, Tianjin University of Science and Technology, Tianjin, P.R. China
ABSTRACT
ARTICLE HISTORY
Received 30 April 2019
Accepted 29 June 2019
The synthesis of the methyl glycyrrhetinate glycosides and
inhibition of a-glucosidase were studied. The carboxyl group
of glycyrrhetinic acid was methylated, and glucose and
galactose were introduced into the hydroxyl group to obtain
compounds 7 and 12. Compound 1, 2, 7, 12 and glycyrrhizic
acid (GL) were evaluated for their inhibitory activities against
a-glucosidase. As a result, Compound 1, 2, 7, 12 and GL all
showed significant a-glucosidase inhibitory activity and IC50
values were 0.465, 1.352, 0.759, 0.687 and 2.085 mM, respect-
ively, and acted as non-competitive inhibitors. The activity
of the compound 2, 7, 12 was lower than compound 1, but
significantly higher than GL. Therefore, it was concluded that
the change of structure in glycyrrhetinic acid by chemical
modification had certain effect on bioactivity, and the change
of carboxyl group, hydroxyl group and the type of mono-
saccharide introduced were the influencing factors.
KEYWORDS
Glycyrrhetinic acid;
monoglycosylation;
a-glucosidase inhibition
CONTACT Zhen-Yuan Zhu
These authors contributed equally to this work.
zhyuanzhu@tust.edu.cn
ꢀ
Supplementary information for this article can be accessed at https://doi.org/10.1080/14786419.2019.1639181.
ß 2019 Informa UK Limited, trading as Taylor & Francis Group

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W. ZHANG ET AL.
1. Introduction
Diabetes mellitus (DM) is a common chronic metabolic disorder characterised by
hyperglycemia. It is now widely recognised that its main hazard is due to complica-
tions caused by long-term hyperglycemia, including hyperglycemia, diabetic ketoacido-
sis and non-ketotic hyperosmolar coma. Serious long-term complications include
cardiovascular disease, chronic renal failure and retinal damage. Therefore, it has
always been considered as the main health killer. DM is mainly divided into insulin-
dependent I (Type I Diabetes mellitus, T1DM) and non-insulin-dependent II (Type II
Diabetes mellitus, T2DM). T1DM accounts for 10% of all DM cases. It occurs mostly in
children and adolescents, and it is caused by the destruction of insulin-producing pan-
creatic islet b cells by the autoimmune system, which leads to decreased insulin pro-
duction. T2DM accounts for 90% of DM cases, and is more common in adults and the
elderly. Current studies suggest that its occurrence and development are closely
related to islet cell damage and insulin resistance, and familial genetic factors also are
significant. With the improvement of people’s living standards and lifestyle changes,
the incidence of T2DM continues to grow and develops toward youthfulness.
According to predictions, the prevalence of T2DM will increase from the current 350
million to 592 million by 2035. Therefore, the prevention and treatment of diabetes
has become an urgent problem to be solved (Wu et al. 2011; Zhang et al. 2006).
One of the drugs for the treatment of T2DM is a-glucosidase inhibitor. These drugs
act on the a-glucosidase in the small intestine mucosa, competitively inhibiting the
activity of this enzyme in the small intestine. The absorption of glucose and the
hydrolysis rate of starch are reduced, and even achieving to inhibit completely,

NATURAL PRODUCT RESEARCH
3
thereby achieving the purpose of reducing postprandial blood glucose (Zhen et al.
2017). a-Glucosidase inhibitors which have been used in clinical treatment include
acarbose, voglibose, miglitol and so on (Picazo et al. 2017).
Glycyrrhetinic acid (GA, compound 1) belongs to pentacyclic triterpenoids, which is nat-
urally present in various licorices (such as Glycyrrhiza glabra, Ural licorice, Yunnan licorice,
etc.) and is one of the main active constituents of licorice. It has a variety of pharmaco-
logical effects, such as anti-cancer (Shen et al. 2017), anti-ulcer (Radwan et al. 2016), anti-
oxidation (Li et al. 2012), anti-inflammatory (Li et al. 2017), and reducing blood lipids
function (Kalaiarasi et al. 2009), etc. Consequently, GA is a leading compound of Chinese
medicine with great research and development potential, and is widely used in medicines
and health foods. Not only that, GA is also added to all kind of cosmetics, in addition to
better whitening effect, but also anti-inflammatory, emollient, scavenging oxygen free rad-
icals and UV protection (Ljiljana et al. 2016; Kong et al. 2015).
Since there are two configurations of hydrogen on C-18 in the GA structure, GA is fur-
ther divided into 18a-GA and 18b-GA. But the naturally occurring 18b-GA is much more
than 18a-GA, so the GA mentioned in the paper is 18b-GA. The previous researchers
firstly protected the C-3 hydroxyl group of GA with acetyl group, and the C-30 carboxyl
group was activated by SOCl2 and then reacted with some aromatic groups, alicyclic
groups, heterocyclic groups and aromatic groups to obtain amides and esters. Comparing
the anti-inflammatory activities of these derivatives, it was found that these derivatives
basically exhibited excellent anti-inflammatory activity, and even some derivatives showed
better anti-inflammatory effects than prednisolone and indomethacin (Radwan et al.
2016). Furthermore, other researchers took derivatives of GA amino phosphonates
as inhibitors of NF-jB pathway and cell proliferation to achieve the anti-tumor effect
(Jin et al. 2018). A large number of literatures have reported the anti-inflammatory,
anti-tumor and anti-cancer activities of GA and its derivatives, but the hypoglycemic
activity of GA and its derivatives have yet to be deeply investigated.
The previous research found that GA and glibenclamide have comparable
hypoglycemic effects in a rat model of hyperglycemia established by streptozotocin
(Kalaiarasi & Pugalendi. 2009).
What’s more, glycyrrhizic acid (GL), its aglycone is GA and its structure also includes
glucuronic acid, reduced blood glucose and HOMA-IR values in type 2 diabetic rats
according to the previous report. It also reduced the level of hyperglycemia, hypergly-
cosylated hemoglobin and serum cholesterol in streptozotocin-induced diabetic rats,
and improved the level of serum insulin, glucose tolerance and the number of islet
cells (Eu et al. 2010).
In the light of above considerations, in this article, some GA derivatives containing
methyl glycyrrhetinate glycosides were carefully designed and obtained. These GA
derivatives were expected to have better a-glucosidase inhibitory activity than GA.
2. Results and discussion
2.1. Chemistry
The general procedures for the synthesis of methyl glycyrrhetinate glycosides are
shown in Scheme 1. In order to prevent the GA C-30 carboxyl group from reacting, it

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W. ZHANG ET AL.
was first protected by ester group (compound 2). The hydroxyl groups of glucose and
galactose were protected by benzoyl groups (compounds 4 and 9) and then treated
with 33% HBr to give the corresponding bromides to react with GA (compounds 5
and 10) (Ruiz et al. 2009). Compounds 5 and 10 were reacted with compound 2 under
catalysis by silver trifluoromethanesulfonate, respectively, to obtain compounds 6 and
11, and finally the benzoyl groups were removed using a sodium methoxide–metha-
nol system to obtain the final product compounds 7 and 12 (Schwarz et al. 2014; Fan
et al. 2016). All compounds were identified by nuclear magnetic resonance spectros-
copy and high resolution mass spectrometry.
1
The signal peaks of –OMe in H NMR (d 3.69) and 13C NMR (d 55.07) of compound
2 were substantially consistent with the previous literature (Zou et al. 2016), indicating
that –COOH in compound 1 had been methylated (Supplementary information Figure
1
S1). In the H NMR of compound 6, d 8.02–7.33 (m, 20, ArH), d 5.67(d, J ¼ 7.8 Hz, 1H,
H-1’) were the signal peaks of benzoyl hydrogen and anomeric hydrogen in glucose,
respectively. In 13C NMR, d 133.56–128.42, d 103.27 were the signal peaks of benzoyl
carbon and anomeric carbon, respectively (Supplementary information Figure S2); d 8.
1
05–7.13 (m, 20, ArH), d 5.89 (d, J ¼ 7.24 Hz, 1H, H-1’) in the H NMR of compound 11
were the signal peaks of benzoyl hydrogen and anomeric hydrogen in galactose. And
d 136.73–124.75, d 102.70 in 13C NMR were the signal peaks of benzoyl carbon and
anomeric carbon in galactose (Supplementary information Figure S4), which fully dem-
onstrated that benzoylated glucose and galactose had reacted with GA successfully.
What’s more, the coupling constants of compounds 6 and 11 anomeric hydrogen
were7.8 Hz, 7.24 Hz, respectively, which proved that the bonds formed were b-type. In
1H NMR and 13C NMR of compounds 7 and 12, the signal peaks of benzoyl group dis-
appeared, indicating that the hydroxy group on glucose and galactose had been
deprotected (Supplementary information Figures S3, S5). The mass of the molecular
peaks of compounds 6, 7, 11, and 12 were 1085.4971, 669.3984, 1085.4996, 669.3979
in the mass spectrums, respectively, and the error did not exceed 5 ppm
(Supplementary information Figure S6).
2.2. a-Glucosidase inhibition
2.2.1. a-Glucosidase inhibitory activity
As described in Supplementary information Figure S7, compound 1, 2, 7, 12 and GL
had certain inhibition activity on a-glucosidase at different concentrations and it was
in a dose-dependent manner (Juncheng et al. 2018). The a-glucosidase inhibitory
activity of these five compounds increased with the increasing sample concentrations
from 0.125 to 4 mg/mL and the inhibition rate tended to be gentle at the concentra-
tion of 2 mg/mL. The half-maximal inhibitory efficiency of inhibitor (IC50) value could
be used to assess inhibitory efficiency of the inhibitor (Hashim et al. 2015). The data in
Supplementary information Table S1 showed that the IC50 value of the acarbose (posi-
tive control) was 0.035 mM and the IC50 values of compounds 1, 2, 7, 12 and GL were
0.465, 1.352, 0.759, 0.687 and 2.085, respectively.
Referring to Supplementary information Figure S7 and Table S1, when the carboxyl
group of GA was esterified (compound 2), the a-glucosidase inhibitory activity was

NATURAL PRODUCT RESEARCH
5
obviously decreased, indicating that the carboxyl group of GA had an important influ-
ence on the a-glucosidase inhibition activity. When the hydroxyl group of GA was
introduced glucose and galactose (compound 7 and 12), the a-glucosidase inhibition
activity decreased, but compared with GL, their inhibition activity was greatly
enhanced. This indicated that the sugar introduced at the C-3 position was different
and the activity was different. The type of monosaccharide introduced was also very
an important factor.
2.2.2. Type of a-glucosidase inhibitory
To clarify the a-glucosidase inhibition mode of compounds 1, 2, 7 and 12,
Lineweaver-Burk plots were generated. As shown in Supplementary information Figure
S8, the rate of enzymatic reaction varies with substrate concentration and was linear.
When the concentration of these four compounds increased from 0.5 to 2 mg/mL, the
slope of the straight line decreased, the y-intercept (1/Vmax) increased, and the x-axis
intercept (1/Km) remained unchanged, indicating that the initial velocity of the max-
imum reaction began to decrease and Km did not change. The reaction process con-
formed to the non-competitive reversible inhibition type, that is, the reversible
inhibition types of a-glucosidase of these four samples were non-competitive.
3. Experimental section
All experimental procedures are described in the supplementary materials.
4. Conclusions
In conclusion, the final products compounds 7 and 12 were obtained by a stepwise
1
reaction of compound 1 with glucose and galactose. Analysis of H NMR and 13C NMR
1
of compounds 7 and 12, d 4.12 ppm and d 4.19 ppm in H NMR were the signals of
glucose and galactose anomeric hydrogen, respectively, d 105.85 ppm and d
105.91 ppm in 13C NMR were the signals of glucose and galactose anomeric carbon,
respectively. The coupling constants of compounds 6 and 11 anomeric hydrogen were
7.8 and 7.24 Hz, respectively. This indicated that glucose and galactose had success-
fully reacted with the hydroxyl group of compound 1 and were linked in b configur-
ation. The a-glucosidase inhibition activity assay illustrated the significant change of
bioactivity in compound 1, 2, 7, 12 and GL. Thus, the conclusion that the change of
structure in GA by chemical modification had certain effect on bioactivity could be
drawn, and the change of carboxyl group, hydroxyl group and the type of monosac-
charide introduced were the influencing factors.
Funding
This work was financially supported by the National Key Project of Research and Development
Plan of China (2016YFD0400803), the National Natural Science Foundation of China (31871791),
the Technology Program of Tianjin, China, (18ZYPTJC00020) and the key program of the
Foundation of Tianjin Educational Committee (2018ZD06).

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W. ZHANG ET AL.
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