Chemical characterization and antimicrobial activity of Baccharis vulneraria Baker essential oil against strains of microorganisms that cause cutaneous infections

Abstract Baccharis vulneraria Baker is used popularly for the treatment of skin infections. So, this study aimed investigate the antimicrobial activity and chemical characterization of the essential oil (EO) against microorganisms that cause cutaneous infections. The EO was analyzed by GC-MS. The antimicrobial test was performed using the serial microdilution method, and the antimicrobial activity was determined by the minimum inhibitory concentration against Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli, Pseudomonas aeruginosa, Candida albicans, Trichophyton interdigitale, Trichophyton rubrum, Fusarium solani and Fusarium oxysporum within the 32-0.0625 mg/mL concentration range. Were identified 31 EO compounds. Its major compounds are bicyclogermacrene, trans-cadin-1,4-diene, β-caryophyllene, and germacrene A. EO showed antifungal action against T. rubrum and T. interdigitale (2 and 4 mg/mL MIC, respectively). The growth of C. albicans, at 4 mg/mL, decreased by 50% compared to control. The oil had no significant potential for other microorganisms at the selected concentrations. Graphical Abstract


Introduction
Baccharis vulneraria Baker, popularly known as Erva-Santa (holy herb), is found in southern and southeastern Brazil.The species is used topically in folk medicine to aid in the healing of skin wounds (Stüker 2007), which can be related to skin infections caused by bacteria and fungi.Among them are Gram-positive bacteria as Staphylococcus spp.; Gram-negative bacteria as Pseudomonas aeruginosa and Escherichia coli; yeasts, mainly Candida spp.; Trichophyton sp., dermatophyte genera more prevalent in worldwide; and non-dermatophyte filamentous fungi as Fusarium sp.(Heidrich 2015;Guevara-Suarez 2016;Clebak and Malone 2018;Denamur 2021).
The essential oil (EO) of the plant was characterized and its antimicrobial potential was evaluated once (Stüker 2007).However, activity against dermatophytes and Fusarium sp. has not been evaluated yet.Thus, the study aimed investigate the antimicrobial activity and chemical characterization of the EO against microorganisms that cause cutaneous infections.

Yield and chemical composition of the essential oil
Based on the methodological procedures described in Supplementary Material (Experimental S1), the EO showed a yield of 0.08%.Paredes et al. (2020), with the EO of species of Baccharis (B. boliviensis, B. densiflora, B. latifolia, B. papillosa, B. tola) achieved yields similar to those of this study, indicating that the species has a low yield of EO and corroborates the present study, in which leaves were collected during the flowering period, which causes a low yield of EO.
Chromatography showed 57 compounds in the EO, out of which 31 were identified (Table S1).Its major compounds are bicyclogermacrene (14.68%), trans-cadin-1,4-diene (13.30%), β-caryophyllene (10.48%), and germacrene A (8.22%), all the sesquiterpene group.Pacheco et al. (2021) point out that sesquiterpenes and monoterpenes are the major constituents found in essential oils, as also reported in the present study, in which sesquiterpenes make up 69.7% of the total compounds.Sesquiterpenes have a favorable function in controlling fungi, bacteria, insects, and other pests, thus explaining the potential of the activities of essential oils in which these compounds are found in greater proportions (Bakkali et al. 2008).
As in our study, Stüker (2007) identified that the oil of B. triplinervium (synonymous of B. vulneraria), collected in the summer and spring, was mostly composed of hydrocarbon sesquiterpenes, in which bicyclogermacrene, germacrene D, trans-caryophyllene and 12,14-labdadien-8-ol corresponded to the main components, with few changes between the samples.Hydrocarbon sesquiterpenes were also the major group of compounds found in our study and the bicyclogermacrene compound was the majority.Therefore, our work corroborates with the only study found in the literature concerning the chemical composition of B. vulneraria and also emphasizes the need to verify the period of collection of the plants for the standardization of the oil.Inhibition of pathogen growth may be related to an important component of the EO of the species, as well as to the activity of its major compounds, or the synergism of all compounds found in them (Borges 2019).

Antimicrobial activity
The evaluation of the ability to inhibit the growth of microorganisms associated with skin infections showed that the activity of the EO varied according to the assessed microorganism, demonstrating an antifungal potential against the dermatophyte fungi Trichophyton rubrum and Trichophyton.interdigitale (MIC = 2 mg/mL and 4 mg/mL, respectively).
The choice to test 32 mg/mL as the highest concentration in this work is based on the use of conventional topic antifungal ketoconazole and miconazole, used in 2% in cream formulations (US FDA 2023), since B. vulneraria is used topically in folk medicine (Stüker 2007).For Fusarium solani and Fusarium oxysporum, Candida albicans, E. coli, P. aeruginosa, Staphylococcus aureus and Staphylococcus epidermidis, the EO had no inhibition potential, showing a MIC higher than 32 mg/mL (Table S2).Nevertheless, for the tested C. albicans strain, 4 mg/mL of EO showed inhibition of at least 50% of the microorganism compared to the growth control (IC50).As the MIC of several antifungals used in the clinic for Candida sp. is determined through the inhibition of 50% (CLSI 2015), the result of this work shows that the tested strain is more susceptible to the oil in comparison with the tested bacteria and Fusarium samples, which presented an IC50 equal to or greater than 16 or greater.Unlike this work, the study of Stüker (2007) showed activity against P. aeruginosa at 5 mg/mL and against C. albicans at 10 mg/mL of the oil of B. vulneraria obtained from plants collected in the summer.Borges (2019), in evaluating new antifungal agents among fourteen species of plants from the Atlantic Forest, found that the EO of the species B. platypoda and B. trinervis was effective against the growth of F. solani at concentrations greater than 0.8 mg/mL of EO.In the present work, B. vulneraria, at 32 mg/mL, was not enough to inhibit 50% of microbial growth.The discrepancy may be due to the different fungal isolates tested.Valarezo et al. (2015), when evaluating the biological activity of the EO of B. obtusifolia Kunth, found results similar to those of this work, observing no action against P. aeruginosa, E. coli and S. aureus.In addition, they found antifungal action against Trichophytum species tested in the present work.The Trichophyton species account for about 94.5% of all dermatophytic infections (Heidrich 2015) and, in this work, the greatest potential of the EO of B. vulneraria was against this microorganism.
The results found in this work confirm the popular use of B. vulneraria to treat skin mycoses, mainly dermatophytosis and candidiasis through topical application (Stüker 2007).The results of this work presented a significant contribution to the identification of biological activities of essential oils from native Brazilian flora.

Conclusion
The essential oil of B. vulneraria obtained in state of Rio Grande do Sul, Brazil, in the summer, consisted mainly of sesquiterpenes, in which Bicyclogermacrene, Trans-cadin-1,4 diene, β-caryophyllene, and Germacrene A were the main compounds.It showed potential for antifungal action against the dermatophyte fungi T. interdigitale and T. rubrum, besides being able to inhibit half of the growth of the tested C. albicans, confirming the popular use of the plant species for the treatment of fungal cutaneous infections.However, the oil did not show antibacterial potential against the evaluated bacteria.