Research Article: Study of the potential adverse effects caused by the dermal application of Dillenia indica L. fruit extract standardized to betulinic acid in rodents

Date Published: May 31, 2019

Publisher: Public Library of Science

Author(s): Flávia S. Fernandes, Gustavo S. da Silva, Alexandre S. Hilel, Ana C. Carvalho, Karina V. T. Remor, Aline D. Schlindwein, Luiz A. Kanis, Daniel F. Martins, Maicon R. Kviecinski, Hans-Joachim Lehmler.


This study aimed to evaluate the potential adverse effects of the dermal administration of Dillenia indica Linnaeus (D. indica) fruit extract in healthy rodents; the extract was standardized to betulinic acid. In the initial phase, the acute effects were evaluated on the skin application site of a single extract dose. A skin irritation test was performed in male Wistar rats (n = 8/group) receiving the extract (50–150 mg/mL) with betulinic acid (0.5–1.5%, respectively). A photosensitivity test was performed in male BALB/c mice (n = 6/group) receiving the extract (150 mg/mL). Afterwards, other BALB/c mice (n = 20, male:female, 1:1) were used to assess the systemic alterations caused by 14 daily repeated doses (150 mg/mL) by monitoring the effects on mortality, body morphology, behavior, nutrition status, neuromotor reactions, organ morphology and weight, and blood tests. At this time, 0.5 mg/mL clobetasol was used as the positive control. The skin irritation index suggested that negligible skin irritation had occurred, even when the extract was applied to the rat skin at 150 mg/mL. However, the extract acted as a photosensitizer on mouse skin, showing a photosensitizing activity close to that of 10 mg/mL 5-methoxypsoralen. Repeated doses caused no mouse mortality, aggressiveness, piloerection, diarrhea, convulsions, neuromotor alterations or nutrition status changes. The mouse organ weights did not change, and the mice did not have alterations in their blood compositions. Clobetasol caused a reduction in the mononuclear leukocyte numbers. In general, the data suggest that the extract was safe in healthy rodents but indicate that caution should be taken with the photosensitizing activity; in addition, this activity should be further explored as it may be useful for phototherapeutic drug development.

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This work was originated from an interest in natural products that could be sources of innovative drugs for the treatment of immune-mediated dermatological disorders, such as psoriasis. Psoriasis is currently the most prevalent autoimmune disease worldwide [1,2]. The available treatment has a low efficacy and many side effects [3,4]. Therefore, a survey was initially carried out with people who worked with folk medicine and integrative therapies. This survey included more than one hundred participants that included elderly and pastoral health groups in the southern region of Brazil. Based on this survey, the species Dillenia indica Linnaeus (D. indica) was chosen to be investigated more thoroughly [5].

The research question approached by this work was based on several previous studies that showed the beneficial bioactivities of D. indica extracts and derivatives. Betulinic acid, a major constituent of D. indica extract, has been extensively studied in recent years. This molecule was mostly studied for its benefits. Most previously conducted toxicological studies on betulinic acid used cytotoxicity assays in cell culture and focused on antitumor activities [54,55].

The extract caused no irritation on the skin of rats; however, it acted as a photosensitizer on the mouse skin. No systemic toxicity was observed from the administration of repeated doses of extract on the intact skin of healthy mice. In general, the data suggest that the use of D. indica fruit extract is safe; this is important for cases where the extract may eventually become a constituent of a future dermal application formulation. This photosensitizing effect deserves to be explored further as it may be interesting for the development of drugs used in the phototherapy of some skin diseases. Although this work allows for some inferences to be made about the safety of the dermal administration of betulinic acid, a fair assessment will depend on further studies with the purified compound and an evaluation of betulinic acid administration on wounded skin, which normally has a compromised barrier function. These experiments are in progress in our laboratory.




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