Boesenbergia rotunda and Its Pinostrobin for Atopic Dermatitis: Dual 5-Lipoxygenase and Cyclooxygenase-2 Inhibitor and Its Mechanistic Study through Steady-State Kinetics and Molecular Modeling

Abstract

Human 5-lipoxygenase (5-LOX) and cyclooxygenase-2 (COX-2) are potential targets for suppressing pruritic skin inflammation in atopic dermatitis (AD). In addition, Staphylococcus aureus colonization and oxidative stress worsen AD skin conditions. We aimed to investigate anti-inflammatory activity, using 5-LOX and COX-2 inhibitions, and the anti-staphylococcal, and antioxidant potentials of several medicinal plants bio-prospected from traditional medicine related to AD pathogenesis. Essential oils and hexane fractions were prepared and analyzed using gas chromatography-mass spectrometry. Boesenbergia rotunda hexane extract displayed anti-Staphylococcus aureus (MIC = 10 µg/mL) and antioxidant activities (IC₅₀ = 557.97 and 2651.67 µg/mL against DPPH and NO radicals, respectively). A major flavonoid, pinostrobin, was further nonchromatographically isolated. Pinostrobin was shown to be a potent 5-LOX inhibitor (IC₅₀ = 0.499 µM) compared to nordihydroguaiaretic acid (NDGA; IC₅₀ = 5.020 µM) and betamethasone dipropionate (BD; IC₅₀ = 2.077 µM) as the first-line of AD treatment. Additionally, pinostrobin inhibited COX-2 (IC₅₀ = 285.67 µM), which was as effective as diclofenac sodium (IC₅₀ = 290.35 µM) and BD (IC₅₀ = 240.09 µM). This kinetic study and molecular modeling showed the mixed-type inhibition of NDGA and pinostrobin against 5-LOX. This study suggests that B. rotunda and its bioactive pinostrobin have promising properties for AD therapy.

Keywords: GC–MS; anti-inflammatory; antibacterial; antioxidant; fingerroot; flavonoid.

Figure 1

Atopic dermatitis is a multi-facet inflammatory skin condition characterized by skin microbiome alteration by colonization of S. aureus, oxidative stress, skin barrier defects, and chronic pruritic inflammation. Several proinflammatory mediators are found to be abundant in AD skin compared to normal skin, including LTB₄, PGD₂, and PGE₂. The suppression of acute skin inflammation and pruritus can be done by targeting 5-LOX and COX-2 pathway; LC: lead compound, FLAP: 5-LOX activating protein.

Figure 2

Determination of (A) total phenolic content (TPC) and (B) total flavonoid content (TFC) values of all tested samples. Hexane extract: ultrasound-assisted extraction (UAE); essential oils (EOs); B. rotunda (BR); C. sinensis (CSD); C. fistula leaves (CFL); C. fistula pods (CFF); M. villosa leaves (MV); D. longan peels (DLP); D. longan seeds (DLS); W. trilobata leaves (WT); and A. vulgaris (AV). Note: The TFC values of UAE DLS and all EOs were not determined because they were inactive at the highest tested concentration (5 mg/mL). One-way ANOVA was performed to compare the mean of each sample in each assay, and different letters indicate the significant difference of the mean.

Figure 3

COX-2 inhibitory activity of B. rotunda hexane extract (UAE BR), essential oils (EOs BR), and pinostrobin compared to two standard drugs; NSAIDs diclofenac sodium and corticosteroid betamethasone dipropionate (BD). One-way ANOVA was performed to compare the mean of each sample in each assay. The same letter (a) indicated that the comparison between groups was not significant.

Figure 4

5-LOX inhibitory activity of B. rotunda hexane extract (UAE BR), essential oils (EOs BR), and pinostrobin compared to standard nordihydroguaiaretic acid (NDGA) and corticosteroid betamethasone dipropionate (BD). One-way ANOVA was performed to compare the mean of each sample in each assay, and different letters indicate the significant difference in the mean.

Figure 5

In vitro steady state kinetics of NDGA and pinostrobin using Michaelis–Menten and Lineweaver–Burk plots; (A) Michaelis–Menten plot of NDGA, (B) Michaelis–Menten plot of pinostrobin, (C) Lineweaver–Burk plot of NDGA, and (D) Lineweaver–Burk plot of pinostrobin.

Figure 6

The predicted binding modes and interactions of (R)- (A,B) and (S)-pinostrobin (C,D) enantiomers with the 5-lipoxygenase binding site. The protein surfaces are rendered with blue and red depicting partially positive and negative charges, and neutral charge for grey areas (A,C). Amino acids predicted to form hydrogen bonding are depicted: His432 (green), Thr364 (yellow), Arg596 (blue), and Gln363 (violet). Residues predicted to form hydrophobic interactions: Phe359 (pink), Pro569 (black), Trp599 (turquoise), and Leu368 (lime green) (B,D).