Phytochemical Profiling, Molecular Docking and ADME Evaluation of Bioactive Constituents of Rice Bran (Oryza sativa L.) Against Human Pancreatic α-Amylase

Rice (Oryza sativa L.) is one of the major cereal crops consumed worldwide. During rice milling, the outer layers of the grain are removed to obtain polished rice. These outer layers, collectively referred to as rice bran, contain a diverse range of nutritional and bioactive constituents. Rice bran contains phenolic acids, flavonoids, tocopherols, tocotrienols, γ-oryzanol, phytosterols and other compounds associated with antioxidant and other biological activities. ^[1,2] Phenolic acids reported in rice bran include ferulic acid, p-coumaric acid, caffeic acid, vanillic acid, gallic acid and related compounds. Flavonoids such as catechin, quercetin and other flavonoid derivatives have also been reported in different rice-bran varieties. Rice bran additionally contains vitamin E derivatives and γ-oryzanol, a mixture of ferulic acid esters of sterols and triterpene alcohols. ^[3] The biological importance of rice bran is partly attributed to its phenolic and flavonoid constituents. Phenolic compounds possess hydroxyl groups capable of donating hydrogen atoms and may therefore contribute to antioxidant activity. Rice bran has been reported to possess considerable antioxidant potential, and its bioactive constituents have been investigated for several biological activities. ^[4,5] α-Amylase is an important digestive enzyme responsible for the hydrolysis of dietary starch into smaller carbohydrate fragments. Inhibition of α-amylase can reduce the rate of starch digestion and may consequently help moderate postprandial carbohydrate availability. Therefore, α-amylase has been widely investigated as a target in the search for potential antidiabetic agents. ^[6] Molecular docking is a computational technique used to predict the preferred orientation of a small molecule within a protein binding site and to estimate its relative binding affinity. AutoDock Vina is widely used for molecular docking and virtual screening because of its computational efficiency and scoring approach. ^[10]

The present investigation combines preliminary phytochemical screening with structure-based molecular docking and ADME prediction to explore selected rice-bran-associated compounds against human pancreatic α-amylase. ^[18] SwissADME provides computational predictions of physicochemical properties, pharmacokinetics, drug-likeness and medicinal-chemistry characteristics and was therefore used to further evaluate the leading docked compounds. ^[12,20]

MATERIALS AND METHODS

Preparation of rice bran extract

The collected rice bran was cleaned to remove foreign matter and dried under suitable conditions. The dried material was powdered and subjected to extraction using ethanol and water in 70:30 ratio by maceration process. The resulting extract was filtered and concentrated under reduced pressure or at a controlled temperature. The concentrated extract was stored in a suitable container until further investigation.

Figure 1. Steps involved in the preparation of Rice bran extract: (a) Rice bran; (b) Washing of rice bran; (c) wet rice bran pulp; (d) Maceration using 70:30 ethanol:water; (e) Filtration; (f) Filtrate; (g) Concentration of filtrate; and (h) Concentrated rice bran extract.

Phytochemical	Test performed	Observation	Result
Alkaloids	Mayer’s test	Pale precipitate	+
Flavonoids	Alkaline reagent	Intense yellow colour	+
Phenolic	Ferric chloride test	Dark blue	+
Tannins	Ferric chloride test	Blue-black	+
Saponins	Froth test	Foam	+
Terpenoids	Salkowski test	Reddish brown colour	+
Glycosides	Kaller-killiani test	No reddish-brown ring	-

Figure 2. Phytochemical screening of rice bran

Figure 3. Three-dimensional structure of human pancreatic α-amylase (PDB ID: 1HNY) used as the target receptor.

Figure 4. Molecular docking and protein–ligand interaction analysis of selected rice-bran phytochemicals with human pancreatic α-amylase (PDB ID: 1HNY): (A) quercetin, (B) epicatechin, (C) catechin, and (D) kaempferol. Each panel presents the three-dimensional docking pose and corresponding two-dimensional interaction diagram of the ligand within the α-amylase binding site.

no predicted blood-brain barrier permeation. They also showed zero Lipinski rule violations. Quercetin and kaempferol were predicted not to be P-glycoprotein substrates, whereas catechin and epicatechin were predicted to be P-glycoprotein substrates. ^[13] These computational findings indicate predicted favorable drug-likeness characteristics under the applied prediction models; however, they should not be interpreted as experimental evidence of oral bioavailability.

DISCUSSION

Rice bran is a chemically diverse plant-derived material containing phenolic acids, flavonoids, tocols, γ-oryzanol and phytosterols. Previous studies have documented ferulic acid, p-coumaric acid, vanillic acid, caffeic acid and other phenolic compounds in rice bran, while flavonoids including catechin and quercetin have also been reported. ^[2,3] The present investigation combined preliminary phytochemical screening with computational evaluation of selected rice-bran-associated compounds. The qualitative phytochemical screening indicated the presence of alkaloids, flavonoids, phenolics, tannins, saponins and terpenoids, whereas glycosides were not detected. These findings provide preliminary evidence for the presence of diverse phytochemical classes in the prepared rice bran extract. Molecular docking against human pancreatic α-amylase showed that flavonoid compounds generally produced stronger predicted binding than the selected phenolic acids. Quercetin and epicatechin produced the most favorable binding affinity of -8.6 kcal/mol, while kaempferol and catechin showed -8.5 kcal/mol. The interaction analysis provides additional structural support for the docking results. The leading compounds interacted with several residues located within or around the α-amylase catalytic region. In particular, Asp197, Glu233 and Asp300 are recognized catalytic residues of human pancreatic α-amylase. ^[9] Quercetin demonstrated interactions involving Tyr62, Gln63, Asp300 and Trp59. Epicatechin demonstrated interactions involving Gln63, Glu233, Asp300 and Trp59. Catechin showed interactions involving Tyr62, Gln63, Asp197, Glu233, Asp300 and Trp59. Kaempferol also demonstrated interactions involving residues around the catalytic binding region. The comparable docking scores of catechin and epicatechin are noteworthy because these compounds possess closely related flavan-3-ol structures. Their hydroxyl-rich structures can provide multiple hydrogen-bonding opportunities, while their aromatic rings can participate in π-related interactions. ADME prediction further indicated that the four leading compounds possessed high predicted gastrointestinal absorption and zero Lipinski violations. Their predicted absence of blood-brain barrier permeation may also be consistent with their relatively polar structures and high topological polar surface areas. However, computational ADME results represent predictions rather than experimental pharmacokinetic measurements. ^[12,13] The present findings therefore provide a preliminary computational basis for considering rice-bran flavonoids as potential α-amylase-interacting compounds. Nevertheless, molecular docking alone cannot establish enzyme inhibition or therapeutic efficacy. The predicted interactions require confirmation through in vitro α-amylase inhibition assays, followed by appropriate experimental validation.

CONCLUSION

The present study investigated selected bioactive constituents associated with rice bran using preliminary phytochemical screening, molecular docking and in silico ADME analysis. Molecular docking against human pancreatic α-amylase identified quercetin and epicatechin as the compounds with the most favorable predicted binding affinities, each showing a predicted binding affinity of −8.6 kcal/mol, followed by kaempferol and catechin at -8.5 kcal/mol. Interaction analysis indicated that the leading compounds formed hydrogen-bonding and aromatic interactions with residues located within the α-amylase binding region, including important catalytic residues such as Asp197, Glu233 and Asp300. SwissADME analysis of the four leading compounds showed high predicted gastrointestinal absorption, no predicted BBB permeation and zero Lipinski rule violations. Overall, the results suggest that rice-bran flavonoids, particularly quercetin, epicatechin, kaempferol and catechin, are promising candidates for further investigation as potential α-amylase-interacting natural compounds. However, the findings are computational and should be validated experimentally through α-amylase inhibition assays and further pharmacological studies.

Declarations

Funding

No external funding was received for this study.

Conflict of Interest

The author declares that there is no conflict of interest associated with this study.

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