Oral and Topical Anti-Inflammatory and Antipyretic Potentialities of Araucaria bidiwillii Shoot Essential Oil and Its Nanoemulsion in Relation to Chemical Composition

Abstract

Different parts of Araucaria bidiwillii (bunya pin) trees, such as nuts, seeds, bark, and shoots, are widely used in cooking, tea, and traditional medicines around the world. The shoots essential oil (EO) has not yet been studied. Herein, the chemical profile of A. bidiwillii shoots EO (ABSEO) was created by GC-MS analysis. Additionally, the in vivo oral and topical anti-inflammatory effect against carrageenan-induced models, as well as antipyretic potentiality of ABSEO and its nanoemulsion were evaluated. Forty-three terpenoid components were identified and categorized as mono- (42.94%), sesqui- (31.66%), and diterpenes (23.74%). The main compounds of the ABSEO were beyerene (20.81%), α-pinene (16.21%), D-limonene (14.22%), germacrene D (6.69%), β-humulene (4.14%), and sabinene (4.12%). The ABSEO and its nanoemulsion exhibited significant inflammation suppression in carrageenan-induced rat paw edema model, in both oral (50 and 100 mg/kg) and topical (5% in soyabean oil) routes, compared to the control and reference drugs groups. All the results demonstrated the significant inflammation reduction via the inflammatory cytokines (IL-1β and IL8), nitrosative (NO), and prostaglandin E2 (PGE2) supported by the histopathological studies and immunohistochemical assessment of MMP-9 and NF-κβ levels in paw tissues. Moreover, the oral administration of ABSEO and its nanoemulsion (50 and 100 mg/kg) exhibited antipyretic activity in rats, demonstrated by the inhibition of hyperthermia induced by intramuscular injection of brewer's yeast. These findings advised that the use of ABSEO and its nanoemulsion against numerous inflammatory and hyperthermia ailments that could be attributed to its active constituents.

Keywords: Araucaria bidiwillii; anti-inflammatory; antipyretic; essential oil; immunohistochemical; inflammatory cytokines; nanoemulsion.

Figure 1

GC–MS chromatogram of ABSEO of shoots of Araucaria bidiwillii cultivated in Egypt. Main components were numbered (1–6); α-Pinene (1), Sabinene (2), d-Limonene (3), β-Humulene (4), Germacrene D (5), and Beyerene (6).

Figure 2

(A) Particle size distribution and (B) zeta analysis of Araucaria bidiwillii EO nanoemulsion (mean diameter = 106 nm).

Figure 3

Effect of oral administration of ABSEO and ABSEO nanoemulsion (50–100 mg/kg) on (A) IL-1β, (B) IL-8, (C) Nitric oxide (NO), and (D) PGE2 RNA expression induced by subplantar injection of carrageenan in the hind left paw of rats as compared to standard drug; diclofenac. Data are expressed as (mean ± SE). Statistical analysis was performed by one-way ANOVA and confirmed by Tukey test. p < 0.05 was assumed to denote statistical significance. a—Significantly different from normal group at p < 0.05. b—Significantly different from positive control group at p < 0.05. c—Significantly different from diclofenac group at p < 0.05.

Figure 4

Effect of oral administration of ABSEO and ABSEO nanoemulsion (50–100 mg/kg) on pyrogenesis induced by intramuscular injection of rats with brewer’s yeast as compared to standard drug; diclofenac and paracetamol. Data are expressed as (mean ± SE). Statistical analysis was performed by one-way ANOVA and confirmed by Tukey test. p < 0.05 was assumed to denote statistical significance.

Figure 5

Effect of topical application of ABSEO (5%) and nanoemulsion (5%) on (A) IL-1β, (B) IL-8, (C) Nitric oxide (NO), and (D) PGE2 RNA expression, induced by intraplantar injection of carrageenan in the hind left paw of rats as compared to standard drug diclofenac. Data are expressed as (mean ± SE). Statistical analysis was performed by one-way ANOVA and confirmed by Tukey test. p < 0.05 was assumed to denote statistical significance. a—Significantly different from normal group at p < 0.05. b—Significantly different from positive control group at p < 0.05. c—Significantly different from diclofenac group at p < 0.05.

Figure 6

Photomicrograph of H&E-stained sections of hind paw of rats. (a) Control rat showing normal histological structure. (b–d) Carrageenan model rat showing; marked edema (Ed) and severe diffuse inflammatory cell infiltration (IF) admixed with many neutrophils, (d) vacuities with early thrombus formation (arrow). (e,f) Oral ABSEO treated carrageenan model rats at low dose (50 mg/kg) (e) and high dose (100 mg/kg) (f) showing moderate degree of decreased inflammatory reaction with still presence of edema (Ed) and infiltrated inflammatory cells (IF). (g,h) Oral ABSEO nanoemulsion treated carrageenan model rats at low dose (50 mg/kg) (g) and high dose (100 mg/kg), (h) showing a dose-related good degree of decreased to scares inflammatory reaction. (i,j) Orally (i) and topically (j) diclofenac treated carrageenan model rats showing edema and severe inflammatory reaction (IF), particularly when topically applied. (k,l) Topically applied ABSEO (5%) (k) and ABSEO nanoemulsion (5%) (l) to carrageenan model rats showing good degree of improvement of paw inflammation, particularly for the ABSEO (5%) application. (m) Topically applied soyabean showing marked inflammatory reaction. (n,o) The scores of inflammatory reactions in different treated groups compared to carrageenan model group. Scoring data are presented as median (max–min) using Kruskal–Wallis test followed by the Mann–Whitney U test. p < 0.05 was assumed to denote statistical significance. a—Significantly different from normal group at p < 0.05. b—Significantly different from positive control group at p < 0.05. c—Significantly different from diclofenac group at p < 0.05.

Figure 7

Immunohistochemical stained sections of rats’ hind paw for MMP-9 expression. (a) Control rat, (b) carrageenan model rats, (c,d) ABSEO orally treated carrageenan model rat at low (50 mg/kg) (c) and high (100 mg/kg) (d) doses, (e,f) ABSEO nanoemulsion orally treated carrageenan model rats at low (50 mg/kg) (e) and high (100 mg/kg) (f) doses. (g,h) Diclofenac treated carrageenan model rat orally (g) and topically (h). (i,j) Topically applied ABSEO (5%) (i) and ABSEO nanoemulsion (5%) (j) to carrageenan model rats. (k) Topically applied soyabean to carrageenan model rats. p < 0.05 was assumed to denote statistical significance.

Figure 8

Immunohistochemical stained sections of rats’ hind paw for NF-κB p65 expression. (a) Control rat, (b) carrageenan model rats, (c,d) ABSEO orally treated carrageenan model rat at low (50 mg/kg), (c) and high (100 mg/kg) (d) doses, (e,f) ABSEO nanoemulsion orally treated carrageenan model rats at low (50 mg/kg) (e) and high (100 mg/kg) (f) doses. (g,h) diclofenac treated carrageenan model rat orally (g) and topically (h). (i,j) Topically applied ABSEO (5%) (i) and ABSEO nanoemulsion (5%) (j) carrageenan model rats. (k) Topically applied soyabean to carrageenan model rats. p < 0.05 was assumed to denote statistical significance.

Figure 9

The quantitative image analysis of both MMP-9 and NF-κB p65 expression in in the oral (A,C) and topical * (B,D) experiments presented as OD of the positive brown color in five microscopic fields using image analysis software (Image J, 1.46a, NIH, USA). a—Significantly different from normal group at p < 0.05. b—Significantly different from positive control group at p < 0.05. c—Significantly different from diclofenac group at p < 0.05.