In Vitro and In Silico Assessments of Curcuminoids and Turmerones from Curcuma longa as Novel Inhibitors of Leishmania infantum Arginase

Abstract

Background/Objectives: The anti-Leishmania potential of Curcuma longa and its derivatives, such as curcuminoids, is well-established, yet their mechanisms of action remain underexplored. This study investigates the inhibitory effects of C. longa extracts and curcumin on Leishmania infantum arginase, a key enzyme in polyamine and trypanothione biosynthesis, and evaluates their antiparasitic activity. Methods: Extracts were prepared via rhizome successive maceration with hexane (HEXCURC), dichloromethane (DCCURC), and ethanol (ETOHCURC) and chemically characterized by a combination of chromatographic and spectrometric methods. The inhibition of recombinant L. infantum arginase (LiARG) was assessed by urea quantification, while molecular docking explored interactions between the main compounds annotated in the extracts and the enzyme's active site. Biological activity was tested against L. infantum promastigotes, intracellular amastigotes, and mammalian cells. Results: LC-MS and GC-MS revealed curcuminoids and turmerones as main compounds annotated in the extracts. DCCURC, HEXCURC, and curcumin showed the strongest LiARG inhibition (IC₅₀ = 10.04, 14.4, and 17.55 μg/mL, respectively). Docking analysis revealed that curcumin, demethoxycurcumin, and bisdemethoxycurcumin bind near the active site, with binding energies of -3.43, -4.14, and -3.99 kcal/mol, respectively. Curcumin demonstrated superior anti-promastigote activity (IC₅₀ = 15.01 μg/mL) and selectivity (SI = 12.7) compared to the extracts. It also significantly reduced amastigote burden in infected macrophages (IC₅₀ = 13.6 μg/mL). Conclusions: This is the first report demonstrating that C. longa extracts and curcumin inhibit LiARG. These findings support curcumin's potential as a lead compound for developing multi-target therapies against leishmaniasis, combining enzyme inhibition with direct antiparasitic effects.

Keywords: ADMET; antileishmanial; curcumin; enzymatic inhibition; molecular docking; turmerone.

Figure 1

Chemical structures of curcumin (1), demethoxycurcumin (2), bisdemethoxycurcumin (3), and turmerones (4–6) in alkaline media.

Figure 2

Binding mode of curcuminoids with LiARG obtained by molecular docking. The superposition of the final docking pose of each inhibitor docked into the active site of LiARG is shown in the center. The three-dimensional model of LiARG is displayed in cartoon representation and colored green. The molecular structures of (a) curcumin (1) (pink), (c) bisdemethoxycurcumin (3) (yellow), and (e) demethoxycurcumin (2) (light pink) are shown in stick representation. Residues that participate in interactions are colored in light green. The 2D diagrams show the interactions between (b) 1, (d) 3, and (f) 2. Hydrogen bonds and ionic interactions are depicted as yellow and blue dashed lines, respectively.

Figure 3

Binding mode of turmerones with LiARG obtained by molecular docking. The superposition of the final docking pose of each inhibitor docked into the active site of LiARG is shown in the center. The three-dimensional model of LiARG is displayed in cartoon representation and colored green. The molecular structures of (a) ar-turmerone (4) (grey), (c) α-turmerone (5) (purple), and (e) β-turmerone (6) (orange) are shown in stick representation. Residues that participate in interactions are colored in light green. The 2D diagrams show the interactions between (b) 4, (d) 5, and (f) 6.

Figure 4

Effect of curcumin and PAT in intracellular amastigotes amounts after 48 h of treatment in L. infantum-infected RAW 264.7 cells. Data are expressed as the mean of two independent experiments, each performed in duplicate. Statistical analysis was conducted using One-Way ANOVA followed by Tukey’s post-test, comparing each treatment with untreated cells (control), where (*) = p < 0.05.

Figure 5

Effects of curcumin (1) and the reference drug control PAT on nitric oxide (NO) production by RAW 264.7 macrophages infected with L. infantum. Data are expressed as the mean of two independent experiments, each performed in duplicate. Statistical analysis was conducted using One-Way ANOVA followed by Tukey’s post-test, comparing each treatment with untreated cells (control), where (*) = p < 0.05.