Zingiber Officinale and Sida Cordifolia as Nature’s Defenders: Exploring their Action Against Inflammation and Parasite

During the past ten years, important progress has been achieved in exploring the pathophysiology of inflammation and the role of free radicals in its development. Reactive oxygen species (ROS), generated through the interaction of free radicals with molecular oxygen, increases abnormally during inflammatory processes. This disrupts the balance between oxidative molecules and the body’s antioxidant defences, leading to oxidative stress. [1], [2] Such stress triggers inflammatory pathways that damage cellular structures. If left unchecked, inflammation conditions, often characterized by granuloma formation and leukocyte infiltration. Plant-derived flavonoids exhibit strong anti-inflammatory effects by effectively neutralizing free radicals and regulating inflammatory responses. [3] Gastrointestinal nematodes have a potentially harmful impact to the productivity of swine and other livestock, leading to considerable economic losses. They hinder efficient livestock production by reducing weight gain, impairing growth rates, decreasing litter sizes, and causing organ condemnations at slaughter, as well as fatalities. [4], [5] Despite their impact, parasitic infections in animals are often underestimated, as clinical symptoms may not always be immediately apparent. For years, parasite control has relied heavily on chemotherapeutic drugs. However, prolonged use and improper management of these medications have led to major challenges, particularly the widespread development of resistance among parasitic strains. [6], [7] Increasing reports indicate that certain parasites affecting sheep, horses, cattle, and pigs have developed resistance to conventional anthelmintics, rendering many commercially available treatments ineffective. This growing resistance, coupled with concerns over drug residues in meat, has sparked interest in alternative, eco-friendly anthelmintic solutions. [8], [9] Medicinal plants have emerged as promising candidates, with numerous studies highlighting their potential in combating gastrointestinal nematodes. [10] The decoction of Sida cordifolia roots and Ginger has been traditionally used for the management of intermittent fever accompanied by chills. In this study, we explore its potential anti- inflammatory, antimicrobial, and anthelmintic properties to provide scientific validation for its traditional use. [11]

MATERIALS AND METHODS:

Preparation of plant root extract:

Decoction method was applied to acquire the extracts of Sida cordifolia and Ginger.

Sida cordifolia and Ginger extraction:

The plant material of Sida cordifolia and ginger was taken and crushed into smaller pieces for better extraction. Add 10 grams of crushed plant material to a pot and pour 80ml of water. Heat the mixture over medium heat, bring to a soft boil. Once boiling, reduce the heat to low and simmer slowly. Simmer until the water is reduced to 1/4^th of the original volume. Carefully take the pot away from heat and strain the liquid using a fine strainer to remove excess. [12]

Anti-Inflammatory Activity:

Egg albumin protein denaturation assay:

Preparation of 1% Egg albumin solution:

1 ml of the translucent portion of egg was added to 100 ml of w/v distilled water and stir thoroughly. Reaction mixture of final volume of 5ml was prepared by combining 0.2ml of 1% egg albumin solution, 2ml of sample extract and standard drug (diclofenac sodium) at varying concentrations and 2.8ml of phosphate buffered saline (pH 7.4). For control, 2ml of distilled water was mixed with 0.2ml of egg albumin solution and 2.8ml of phosphate buffered saline, maintaining the total volume at 5ml. Both the sample and control mixtures were incubated at 37^OC for 30 minutes, followed by heating at 70^oC in a water bath for 15 minutes. After cooling, absorbance readings were taken at 280nm using UV VIS spectrophotometer with distilled water serving as blank. The %inhibition of protein denaturation was calculated using the standard formula:

% inhibition = Absorbance of control – Absorbance of test sample / Absorbance of control* 100 [13]

Anthelmintic Activity:

An in-vitro assay was performed to evaluate the anthelmintic efficacy of both Sida cordifolia and Ginger extracts using the earthworm Pheretima Posthuma as a biological model.

Preparation of test sample:

For the in-vitro analysis, samples were formulated by dissolving and suspending various amounts of both extracts in distilled water. The resulting solutions were prepared at concentrations of 50, 100, 250 mg/ml. [14]

Anthelmintic assay:

Indian earthworms, Pheretima posthuma (Annelida), were obtained from waterlogged soil regions, with an average length of 6–8 cm. To eliminate any adhering dirt, they were thoroughly washed with tap water. The worms used in the study measured approximately 1–1.5 cm in size. The anthelmintic evaluation was conducted following the method described by Ajayieoba et al., with slight modifications. Freshly prepared test and reference pharmaceutical served in conducting the assay. The paralysis time was recorded when no movement was observed, except upon vigorous shaking. The time of death was confirmed when the worms showed no response to vigorous shaking or immersion in warm water at 50°C. Albendazole (10 mg/ml) served as the reference standard, while distilled water acted as the designated vehicle control. Each experiment was conducted thrice for accuracy. [15]

Image no: 1 Anthelmintic assay

Statistical Analysis:

The statistical analysis in the anthelmintic assay was performed to ensure the reliability and significance of the results. The data, including duration of immobilization and fatality of Pheretima posthuma (earthworms) at different extract concentrations (50, 100, and 250 mg/mL), were expressed as Mean ± SEM (Standard Error of Mean). To compare the effectiveness of Zingiber officinale, Sida cordifolia, and their combination alongside the reference compound (Albendazole), one-way ANOVA (Analysis of Variance) was employed, followed by Dunnett’s test. This statistical method was used to determine notable variation among the treatment groups and the control. A p-value of less than 0.05 (p < 0.05) was considered statistically significant, indicating that the observed differences in duration of immobilization and fatality were not due to random variation but rather to the effects of the plant extracts. This statistical approach validates the anthelmintic efficacy of the extracts, confirming their potential as natural alternatives to synthetic drugs.

RESULTS AND DISCUSSION:

Anti-Inflammatory Assay:

The anti-inflammatory potential of Sida cordifolia, ginger, and their combination was assessed using the egg albumin denaturation assay. The plant extracts demonstrated a dose-dependent separation of protein structure disruption. Among the tested samples: Ginger extract exhibited greater inhibition of protein denaturation compared to Sida cordifolia across all tested concentrations. The combination of Sida cordifolia and ginger (1:1) showed a synergistic effect, with higher inhibition percentages than either extract alone, suggesting that the compounds complement each other’s mechanisms of action. Diclofenac sodium (standard drug) displayed the highest inhibition, but the combined extract approached its efficacy at higher concentrations. The results indicate that the bioactive compounds in both extracts work together to prevent protein denaturation, reducing the inflammatory response. The enhanced inhibition by the combined extract highlights a potential synergistic interaction between the two plants.

Figure 1. The plant extracts of Sida cordifolia, ginger and their mixture (1:1) along with the reference medication Diclofenac sodium, demonstrated a concentration vs % inhibition curve for anti-inflammatory activity. Absorbance was measured at 280 nm using a UV double-beam spectrophotometer.

Anthelmintic Assay:

The anthelmintic efficacy of Sida cordifolia, ginger, and their mixture was evaluated using Pheretima posthuma earthworms. The onset of paralysis and death was recorded at three different concentrations (50, 100, and 250 mg/ml): Ginger extract induced faster paralysis and death compared to Sida cordifolia alone, suggesting a stronger neuromuscular blocking effect. The combined extract demonstrated enhanced efficacy, especially at 100 mg/ml, where the time to paralysis (15.89 ± 1.7 min) and death (25.31 ± 1.1 min) was comparable to the standard drug Albendazole (16.44 ± 1.8 min and 25.87 ± 2.0 min respectively). At the highest concentration (250 mg/ml), both extracts significantly reduced the time required for paralysis and death, confirming a dose-dependent response. The superior performance of the combined extract indicates a synergistic effect, possibly due to the complementary actions of the active compounds in disrupting the parasites’ neuromuscular functions. These results suggest that the Sida cordifolia and ginger mixture could be a promising natural alternative to synthetic anthelmintics. The mean and SEM were statistically analysed using ANOVA, followed by Dunnett’s test, with P<0.05 considered significant.

Table No: 1. Laboratory assessment of the anthelmintic activity of Sida cordifolia, ginger and their mixture along with Albendazole as standard and distilled water as control

Results are expressed as mean ± SEM, N=3, *P <0.05 as compared to standard, ns= not significant

Figure 2: In vitro evaluation of Anthelmintic activity of Sida cordifolia, ginger and their mixture

Figure 3: In vitro evaluation of Anthelmintic activity of Sida cordifolia, ginger and their mixture