Acute Dermal Irritation/Corrosion Testing in Rabbits of Rhinacanthus nasutus Root Extract Gel Containing 0.2% rhinacanthin C+

Abstract

Introduction and Objective: Rhinacanthus nasutus (L.) Kurz, commonly known as snake jasmine or Thong–Phan–Chang in Thai, is a shrub in the Acanthaceae family. This plant is widely distributed across tropical regions, particularly in Southeast Asia, South China, and India. In Thai traditional medicine, both the leaves and roots of R. nasutus have been used to treat various skin diseases, including eczema, ringworm, and tinea versicolor. The plant exhibits potent antifungal activity against dermatophytes, especially the root, which contains rhinacanthin C, a naphthoquinone compound, as the major antifungal substance. In previous studies, a root extract gel from R. nasutus was developed as a topical herbal formulation for treating skin conditions caused by fungal infections. The 0.1% rhinacanthin C gel preparation has been extensively studied, with empirical data supporting its safety in both animal models and humans. However, the safety profile of a higher–concentration formulation, specifically the 0.2% rhinacanthin C gel, has yet to be thoroughly assessed. This study aimed to evaluate the dermal irritation and corrosion responses of the 0.2% rhinacanthin C gel derived from R. nasutus root extract in rabbits to obtain preliminary safety data of the formulation prior to conducting a dose–response study in patients.

Methodology: The dried roots of R. nasutus were extracted, and the rhinacanthin C content was quantified using reverse–phase high–performance liquid chromatography (RP–HPLC).  The R. nasutus root extract was then used as the active ingredient to prepare a 0.2% rhinacanthin C gel. The quality of the gel was later evaluated, including the gel's physical characteristics, the percentage of rhinacanthin C relative to the labeled amount, microbial and heavy metal contamination, and pH, to ensure its suitability for dermal application. The prepared 0.2% rhinacanthin C gel was then subjected to dermal irritation and corrosion testing in accordance with OECD Guideline 404 (OECD Guideline for Testing of Chemicals: Acute Dermal Irritation/Corrosion). New Zealand White rabbits were used as test animals. The testing process began with the skin corrosion test (initial test), conducted on a single rabbit to observe the dermal response, including erythema and oedema, and to monitor for any signs of skin damage or other adverse reactions. If no corrosion was observed in the initial test, a skin irritation test (confirmatory test) was conducted on two additional rabbits to assess potential irritation further.

Results: The prepared R. nasutus root extract gel (0.2% Rhinacanthin C) exhibited appropriate physical characteristics and contained rhinacanthin C within the specified concentration (90.0–110.0% of the labeled amount). No microbial contamination was detected, and heavy metal contamination did not exceed the limits of Thai Herbal Pharmacopoeia. The formulation had a pH of 5.5. These results confirmed that the gel preparation was suitable for testing acute dermal irritation/corrosion. In the skin corrosion test, no signs of skin damage were observed, indicating that the R. nasutus root extract gel (0.2% rhinacanthin C) did not induce skin corrosion. The skin irritation test revealed mean values of 0.67 – 1.00 for erythema and 0 – 0.67 for oedema, both of which met the no irritation requirement (0 to <1.5 non–irritant). These findings suggest that the 0.2% rhinacanthin C gel did not exhibit dermal irritation.

Discussion: Although the 0.2% rhinacanthin C gel did not exhibit significant skin irritation or corrosion, it is important to note that the mean values of erythema and oedema scores were slightly higher than those observed in our previous study using the 0.1% rhinacanthin C gel, which showed mean values of 0.33–0.67 for erythema and 0 for oedema. This suggested that formulations containing rhinacanthin C at concentrations above 0.2% might pose an elevated risk of skin irritation.

Conclusion and Recommandation: The R. nasutus root extract gel containing 0.2% rhinacanthin C demonstrated no significant skin corrosion or irritation in New Zealand White rabbits in accordance with OECD Guideline No. 404. These findings suggest that the 0.2% rhinacanthin C gel preparation is safe for topical application and is suitable for further clinical trials to evaluate the efficacy and safety in patients with dermatophytosis.

Key words Rhinacanthus nasutus root extract gel, rhinacanthin C, safety, irritation, corrosion