Evaluation of Bioactive Metabolites and Antioxidant-Rich Extracts of Amaranths with Possible Role in Pancreatic Lipase Interaction: In Silico and In Vitro Studies

Abstract

Fat/carbohydrate-rich diet consumption or elevated secretion of pancreatic lipase (PL) in pancreatic injury results in increased fat digestion and storage. Several metabolites in plant-based diets can help achieve the requirements of nutrition and fitness together. Presently, nutritional metabolites from Amaranthus tricolor, A. viridis, and Achyranthes aspera were assessed and predicted for daily intake. The volatile-metabolite profiling of their extracts using GC-MS revealed various antioxidant and bioactive components. The implication of these specialized components and antioxidant-rich extracts (EC₅₀ free radical scavenging: 34.1 ± 1.5 to 166.3 ± 14.2 µg/mL; FRAP values: 12.1 ± 1.0 to 34.0 ± 2.0 µg Trolox Equivalent/mg) in lipolysis regulation by means of interaction with PL was checked by in silico docking (Betahistine and vitamins: ΔGbind -2.3 to -4.4 kcal/mol) and in vitro fluorescence quenching. Out of the various compounds and extracts tested, Betahistine, ATRA and AVLA showed better quenching the PL fluorescence. The identification of potential extracts as source of functional components contributing to nutrition and fat regulation can be improved through such study.

Keywords: Amaranthaceae; antioxidants; fluorescence quenching; molecular docking; pancreatic lipase; phytochemicals; proximate contents; volatile metabolites.

Figure 1

Total Phenolic Contents (TPC) (as microgram Gallic acid equivalents/milligram dry extracts: µg GAE/mg DE) and Total flavonoid contents (TFC) (as µg Quercetin equivalents/mg DE: µg QE/mg DE) of acetone and methanolic extracts of Achyranthes aspera L. (AA), Amaranthus viridis L. (AV), and Amaranthus tricolor L. (AT) plants parts. (a) TPC of extracts; (b) TFC extracts where, ATL = AT leaves; ATS = AT stems; ATR = AT roots; AVL = AV leaves; AVS = AV stems; AVR = AV roots; AAL = AA leaves; and AAS = AA stems. Values are significant at p ≤ 0.01 and ≤ 0.05 (Different alphabets and roman numerals on the bars, show statistical difference in values within the same group).

Figure 2

Antioxidant potential of extracts of Achyranthes aspera L. (AA), Amaranthus viridis L. (AV), and Amaranthus tricolor L. (AT). (a) Extracts vs. EC₅₀ (µg/mL) in DPPH assays; (b) Extracts vs. EC₅₀ (µg/mL) in ABTS assays; (c) FRAP values of extracts where, A = ascorbic acid; ATLA = acetone AT leaves; ATSA = acetone AT stems; ATRA = acetone AT roots; ATLM = methanolic AT leaves; AVLA = acetone AV leaves; AVSA = acetone AV stems; AVLM = methanolic AV leaves; AALM = methanolic AA leaves; T = trolox. Values are significant at p ≤ 0.01 and ≤ 0.05 (Different alphabets on the bars, show statistical difference within the same group).

Figure 3

3D orientation of the docked conformations of 1ETH with ligands. (a) Betahistine; (b) α-Tocopherol; (c) γ-Tocopherol; (d) Tocopheryl acetate; and (e) Phytonadione.

Figure 4

Fluorescence spectra of porcine pancreatic lipase (PPL) shows fluorescence quenching in the presence of extracts and compounds. Fluorescence intensities (FI) vs. wavelengths (λ nm) at different temperatures (a) 310; (b) 320; and (c) 330 K temperatures (PPL + extracts: 20, 100, and 200 mg/L); Insets show: PPL + orlistat: 1.0, 2.0, and 3.0 µM/L (a), PPL + betahistine: 0.4, 0.6, and 0.8 µM/L (b) and Comparative spectra of 3 concentrations of AALM, ATRA, and AVLA (c), at 310 K.

Figure 4

Fluorescence spectra of porcine pancreatic lipase (PPL) shows fluorescence quenching in the presence of extracts and compounds. Fluorescence intensities (FI) vs. wavelengths (λ nm) at different temperatures (a) 310; (b) 320; and (c) 330 K temperatures (PPL + extracts: 20, 100, and 200 mg/L); Insets show: PPL + orlistat: 1.0, 2.0, and 3.0 µM/L (a), PPL + betahistine: 0.4, 0.6, and 0.8 µM/L (b) and Comparative spectra of 3 concentrations of AALM, ATRA, and AVLA (c), at 310 K.

Figure 5

Kinetics of PPL fluorescence quenching: Stern-Volmer plots show F₀/F vs. concentrations [Q].

Figure 6

Kinetics of PPL fluorescence quenching: Plot shows relationship between interaction (K_a) of extracts and temperature (lnK_a vs. 1/T).