Banca de DEFESA: EMESON FARIAS ARAUJO SANTOS

Uma banca de DEFESA de MESTRADO foi cadastrada pelo programa.
STUDENT : EMESON FARIAS ARAUJO SANTOS
DATE: 20/11/2023
TIME: 09:00
LOCAL: Google Meet
TITLE:

Evaluation of curcumin derivatives for biotechnological applications: inhibition of carbohydrate digestive enzymes, antioxidant capacity and biophysical studies.

KEY WORDS:

Diabetes mellitus; Enzymology; Enzyme inhibition; Antihyperglycemic.

PAGES: 60
BIG AREA: Ciências Exatas e da Terra
AREA: Química
SUMMARY:

The use of digestive enzyme inhibitors is one of the alternatives applied both as a therapeutic target in treating diabetes mellitus and antiviral agents, as well as agricultural applications in pest control. The inhibitory potential of curcumin derivatives (natural products) on the action of enzymes such as α-amylase and α-glucosidase has been reported, making it important to evaluate new compounds. Therefore, eleven compounds derived from curcumin were characterized by their action as α-amylase and αglucosidase inhibitors. The determination of inhibition of α-amylase activity was performed by the determination of reducing sugars with 3,5-dinitrosalicylic acid (DNS), using acarbose as a positive control. The inhibition of α-glucosidase activity was determined using the method by Tanruean et al. (2019). Antioxidant activity studies of the compounds were carried out using the DPPH• radical scavenging assay and the ABTS•+ radical scavenging assay as experimental procedures. In addition, protein-ligand interaction studies using molecular fluorescence and UV-vis methodology were investigated. The use of Tris-HCl buffer with CaCl2 (pH 7) resulted in a low AC concentration to inhibit 50% of the enzyme activity (IC50), equivalent to 19.1 µM, while in phosphate buffer (pH 7) it obtained if IC50 = 29.8 µM. Furthermore, increasing the temperature during the enzyme and starch incubation time led to lower IC50 values. A screening assay using 120 µM for all compounds was performed to verify their potential to inhibit α-amylase. In this assay, only four derivatives showed inhibition greater than 50%, and of these, only 6 had an inhibitory potential similar to AC. In the α-glucosidase activity screening assay, applying a single dose of 250 µM of the compound against a concentration of 0.02 U mL-1 of the enzyme, it was possible to verify that only six compounds expressed inhibitory activity above 50%, which compounds 1, 4, 6, 7, 8 and 10 obtained the following IC50 values 37.37 (± 1.12), 24.53(± 1.39), 24.93 (± 0.73), 17.67 ( ± 0.58), 19.21( ± 0.43), 27.59 (± 0.38) while the standard expressed 328 (± 6) µM. Based on classical kinetics studies, it was possible to characterize the type of inhibitor applied to the enzymes, obtaining mixed inhibition as a result, using compound 6 on α-amylase and compound 7 on α-glucosidase. For classical studies of protein-ligand interaction using molecular fluorescence methodology, compound 6 was used. Therefore, increasing temperature (22 to 38ºC) versus inversely proportional values of KSV (7.75 to 5.35×104 M-1 ) characterizes an indicative profile of the static quenching process. In which it was confirmed from the values of the biomolecular extinction rate constant (Kq) with the values (5.35 to 7.75×1012 M-1 s -1 ), with values greater than the limiting diffusion rate constant of biomolecules 2.0×1010 M-1 s - 1 is indicative of static quenching. In addition to the binding parameters, thermodynamic parameters were investigated in order to understand the main intermolecular forms present in the α-amylase complex with compound 6. The main intermolecular interactions between α-amylase and compound 6 are hydrogen bonds and Van der Waals forces. In addition, it was possible to verify that the interaction process between the enzyme and the ligand was thermodynamically spontaneous, considering that the G < 0. From the evaluation of conformational changes of αamylase through three-dimensional fluorescence, there was a change in its three-dimensional structure. This is because the addition of compound 6 led to changes in the microenvironment of aromatic amino acids, given that there was a reduction in the fluorescence signal in peaks 2 and 3. Finally, the quenching mechanism, preferably the static one, was confirmed by monitoring the ground state of the protein through UV-vis, at which the absorbance value of the complex is different from the sum of the absorption of the enzyme plus the compound.

COMMITTEE MEMBERS:
Interno(a) - 1006306 - JADRIANE DE ALMEIDA XAVIER
Presidente - 1613338 - JOSUE CARINHANHA CALDAS SANTOS
Externo(a) à Instituição - LUZIA VALENTINA MODOLO - UFMG