Prediction of anti-diabetic activity of flavonoids targeting α-glucosidase - PowerPoint Presentation

Slide1

Prediction of anti-diabetic activity of flavonoids targeting α-glucosidase

Marisa Freitas, Adelaide Sousa, Daniela Ribeiro, Eduarda Fernandes

Slide2

Introduction

Diabetes mellitus is a pandemic disease and is one of the main threats to human health.

Figure 1.

Global projection for diabetes epidemic: 2011-2030:

source

Reference: Ansari et al. 2015. International Journal of Diabetes Research 4(1): 7-12

Slide3

Introduction

GLUT4: Glucose

transporter

Pathophysiology

Adapted

from

: https

://www.pinterest.pt/pin/161707442848089173/

Slide4

Introduction

Source

: http

://www.giostar.com/Therapy/diabetes-type-2/

Hyperglicaemia

Slide5

Pharmacological treatment

Insulin

secretagogues

Biguanides

Thiazolidinediones

α

-

glucosidase

Sulfonylureas

Meglitidines

Tolbutamide

Chloroprapamide

Glyburide

Repaglinide

Metmorfin

Rosiglitazone

Acarbose

Miglitol

Introduction

Slide6

Introduction

Sucrase

-isomaltase

Maltase-glucoamylase

Glucose

Starch

α

-

amylase

Maltose

α

-

glucosidase

Slide7

Source

:

Adapted

from

http://www.servier.com/Powerpoint-image-bank

12

α

-

Glucosidase

Carbohydrates

Glicose

Acarbose

SGLT1

Intestinal

villy

Enterocyte

Without

acarbose

With

acarbose

α

-Glucosidase

Slide8

Side

effects

of

anti-diabetic

drugs

Flatulence

;

Abdominal

discomfort

;

Hepatic

disturbances

;

Contraindicated

in

patients

with

inflammatory diseasesIntroduction

Slide9

Introduction

Flavone

Isoflavone

Flavanone

Flavanol

Flavonol

Anthocyanin

Flavonoids

Slide10

12

Introduction

Inhibition

of

α

-

glucosidase

Biological

activities

of

flavonoids

Slide11

Introduction

a

Slide12

Introduction

a

Experimental

variables

Structure

activity

relationship

SAR

Slide13

AIM

a

1

2

3

Slide14

Methods

a

Chemical structures of the tested flavonoids

Slide15

Methods

a

Chemical structures of the tested flavonoids

Slide16

Methods

a

Inhibitor

Abs

. 405

nm

Slide17

Methods

a

Abs

= 405

nm

Optimization of the

microanalysis

tecnhique

DMSO

Phosphate

buffer, pH=6.8

α-

glucosidase

from

Saccharomyces

cerevisiae

(0 – 0.4 U/mL)

5 min, 37˚C

pNPG

(150 – 2400 µM)

40 - 60 min, 37˚C

Slide18

R

esults

a

Variation

of

α-

glucosidase

concentration



pNPG



= 600

µM

Slide19

R

esults

a

Variation

of

substrate

concentration



α

-

glucosidase



= 0.050 U/

mL

Slide20

Methods

a

Abs

= 405

nm

Study

of

the

inhibitory

activity

of

a

panel

of

flavonoids

Flavonoids

/

Acarbose

/DMSO

Phosphate

buffer, pH=6.8

α-

glucosidase

from

Saccharomyces

cerevisiae

(0.05

U/

mL

)

5 min, 37˚C

pNPG

(600 µM)

30 min, 37˚C

Slide21

Results

a

Compound

Structure

R

2’

R

3

R

3’

R

4’

R

6

R

7

R

8

IC

50

(μM)

A1

(

Flavone

)

-

H

H

H

-

-

-

<20%*

200

μM

a

A2

-

H

OH

H

-

-

-

<20%*

200

μM

aA3-HHOH---<20%* 200 μM aA4-HOHOH---32 ± 4%* 200 μM aA5-OHOHOH---54 ± 3A6-OHOMeOMe---<20%* 100 μM aB1--HH---<20%* 200 μM aB2--OHH---31 ± 4%* 200 μM aB3--HOH---66 ± 2B4--OHOH---66 ± 7a - Inhibitory activity (mean ± SEM %) at the highest tested concentration (in superscript).Table 1. Structures and in vitro α-glucosidase inhibition by the studied flavonoids (IC50 µM, mean ± SEM).

Slide22

Results

Compound

Structure

R

2’

R

3

R

3’

R

4’

R

6

R

7

R

8

IC

50

(μM)

C1

-

H

H

H

-

OH

H

<20%*

200 μM a

C2

-

H

OH

H

-

OH

H

53 ± 4

C3

-

H

H

OH

-

OH

H

≈ 200C4-OHOHH-OHH42 ± 4C5-OHHOH-OHH96 ± 10C6-HOHOH-OHH95 ± 7C7-OHOHOH-OHOH7.6 ± 0.4C8-OHOMeH-OMeH22 ± 2%* 100 μM aC9-OHHOMe-OMeH<20%* 200 μM aC10-OHOHOH-OMeOMe86 ± 6C11-OHOMeOMe-OHOH31 ± 3%* 200 μM aC12-OHOBnOBn-OMeOMe<20%* 200 μM aC13-OHOMeOMe-OBnOBn32 ± 3%* 200 μM aTable 1. Structures and in vitro α-glucosidase inhibition by the studied flavonoids (IC50 µM, mean ± SEM).a - Inhibitory activity (mean ± SEM %) at the highest tested concentration (in superscript).

Slide23

Results

a

Compound

Structure

R

2’

R

3

R

3’

R

4’

R

6

R

7

R

8

IC

50

(μM)

D1

(

Chrysin

)

H

H

H

H

H

-

H

<20%*

50 μM a

D2

(Galangin)

H

OH

H

H

H

-

H

21 ± 3%*

200

μM

aD3(Baicalein)H

HHHOH-H44 ± 3D4HHOHHH-H89 ± 3D5(Apigenin)HHHOHH-H82 ± 6D6(Kaempferol)HOHHOHH-H32 ± 3D7(Luteolin)HHOHOHH-H46 ± 6D8 (Quercetin)HOHOHOHH-H15 ± 3D9(Morin)OHOHHOHH-H32 ± 2Table 1. Structures and in vitro α-glucosidase inhibition by the studied flavonoids (IC50 µM, mean ± SEM).

Slide24

Results

a

Compound

Structure

R

2’

R

3

R

3’

R

4’

R

6

R

7

R

8

IC

50

(

μM

)

E1

(Naringenin)

-

H

H

-

-

-

-

45 ± 3%*

200 μM a

E2

(Eriodictyol)

-

H

OH

-

-

-

-

35 ± 4%*

200 μM a

E3

(Taxifolin)

-OHOH----≈200Positive control:Acarbose-------607 ± 56Table 1. Structures and in vitro α-glucosidase inhibition by the studied flavonoids (IC50 µM, mean ± SEM).a - Inhibitory activity (mean ± SEM %) at the highest tested concentration (in superscript).

Slide25

Results

a

A5

C7

D8 (

quercetin

)

The

most

active

flavonoids

:





Slide26

Results

a

Molecular

docking

calculations

Slide27

Methods

a

Inhibition

type

of

the

most

active

flavonoids



Enzyme



: 0.05 U/

mL



pNPG

:

300, 600 e 1200

µM

Michaelis-

Menten

Equation

:

Lineweaver-Burk

Equation

:

 

Slide28

Methods

a

Inhibition

type

of

the

most

active

flavonoids

Slide29

Results

a

Mixed

Inhibition

Slide30

Results

a

Competitive

Inhibition

Slide31

Results

a

Non-

Competitive

Inhibition

Slide32

Results

a

Flavonoid

Type of Inhibition

Ki (μM)

A5

Mixed

41.0

B3

Mixed

127.0

C7

Competitive

6.5

D8 (quercetin)

Competitive

6.8

E3 (taxifolin)

Non-competitive

347.1

Positive control:

Acarbose

Competitive

457.3

Table 2.

Ki values

(

lM

) for the inhibition of yeast a-glucosidase by the

selected

flavonoids

.

Slide33

Conclusions

a

A

microanalysis

tecnhique

was

implemented

for

the

evaluationof

the

inhibitory

effect

of

flavonoids

against

α

-

glucosidase

.

Enzyme

concentration

: 0.050 U/

mL

Substrate

concentration

: 600 µM

Kinetic

time: 30

minutes

The

substitution

pattern of flavonoids significatively affects their inhibitory activity. The flavonoid structure, the position and number of OH groups are determinant factors for the intended effect.

Slide34

a

Slide35

Aknowlgements

a