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Article

Potential of Essential Oil from Siparuna guianensis A. DC. (Siparunaceae) as an Antimicrobial Adjuvant in Topical Formulations

by
Érica Luiz dos Santos
,
Juliana Mendes Franco Siqueira
,
Genilson da Silva de Jesus
,
Ana Camila Micheletti
and
Nídia Cristiane Yoshida
*
Instituto de Química, Universidade Federal de Mato Grosso do Sul, Campo Grande 79074-460, Mato Grosso do Sul, Brazil
*
Author to whom correspondence should be addressed.
Cosmetics 2025, 12(2), 81; https://doi.org/10.3390/cosmetics12020081
Submission received: 4 February 2025 / Revised: 8 April 2025 / Accepted: 9 April 2025 / Published: 15 April 2025
(This article belongs to the Section Cosmetic Formulations)
Browse Figure
Versions Notes

Abstract

:
Essential oils have enormous versatility as sources of natural fragrances and as active agents in the cosmetic industry. Therefore, the chemical composition and antimicrobial and antioxidant activities of the essential oil from the fresh leaves of Siparuna guianensis A. DC. for cosmetic purposes were analyzed. The GC/MS technique was used to analyze the essential oil and the major constituents found were the sesquiterpenes bicyclogermacrene (32.52%), germacrene D (21.60%), and germacrene B (6.84%) and the monoterpene myrcene (3.66%). The antioxidant activity of the essential oil was evaluated using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical sequestering method and the assay based on the oxidation of 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS). The antioxidant potential of the essential oil was not evidenced in both tests. In vitro tests showed that the studied essential oil, when combined with the antibiotic ampicillin, demonstrated a synergistic effect against clinically resistant Staphylococcus aureus and XDR S. epidermidis strains and an additive effect against S. pseudointermedius and MDR S. epidermidis. On the other hand, the combination of essential oil with gentamicin resulted in synergism when tested against S. epidermidis and an additive effect when evaluated against XDR S. epidermidis. Topical products formulated on the basis of these results exhibited activity against resistant S. epidermidis, demonstrating that the essential oil can act as a valuable ingredient to restore the efficacy of antibiotics against multidrug-resistant bacteria, in addition to improving the olfactory characteristics of the final product.

1. Introduction

In recent years, the use of cosmetics based on plant species has expanded exponentially. They can contain various bioactive molecules that stimulate skin cell regeneration and protect them from cutaneous aging, among other benefits [1,2]. Although cosmetic products are generally not associated with serious health risks, the bioaccumulation of synthetic substances present in their formulations, such as phthalates used to maintain fragrance, dissolve and stabilize essential oils and other aromatic components [3], parabens used as preservatives [4,5], and dioxanes present as emulsifiers and dispersants agents [6], over time has been associated with reproductive disorders, cancer, dermatitis, hair loss, lung damage, and skin aging [2,7,8]. Therefore, despite the potential of Brazilian plant species for wide application in clinical fields [1], which can serve as a source of natural substances, their uses are still underexplored in the development of cosmetic formulations and personal care products.
Plants as a source of active principles for cosmetics offer advantages over synthetic ingredients because they can be used as fixatives, emulsions, antimicrobial, preservatives, or even as multifunctional components [9]. For example, the pharmaceutical and cosmetic industries have increased interest in replacing synthetic antimicrobial agents and antioxidants in dermatological formulations for natural substances [10,11,12]. According to the World Health Organization report, microbial resistance to antibiotics has become a very serious health problem and even a minor infection can be life-threatening. Moreover, products such as cosmetics, for example, may present microbiological contamination both in the production phase and in the product use phase, representing danger to consumers [1,13]. However, to increase the safety and durability of products, the cosmetic industry has sought effective antimicrobial agents of natural origin that have low toxicity, low production costs, and wide application in various areas [13,14,15]. Furthermore, the antimicrobial property of the natural products can be explored alone and/or in combination with antibiotics, both acting as antimicrobial or as an adjuvant, improving or restoring the antimicrobial potential of drugs or topical products, including those that have lost activity against MDR bacteria [13,14]. Additionally, the interaction of UV radiation with the skin surface can result in free radicals or reactive oxygen species (ROS), leading to biochemical transformations in collagen, elastin, and connective tissues. These transformations can affect the firmness and elasticity of the skin, making it dry and wrinkled in appearance [11,12,16]. Antioxidant substances, however, scavenge free radicals and slow down the oxidation process. Moreover, antioxidants with a high reactivity capacity can be used to eliminate ROS [12,16]. Nevertheless, the instability of most antioxidants can cause numerous issues in cosmetic formulations [12]. Antioxidants derived from plants have been highlighted because they contain a mixture of natural substances that can have synergistic effects and thus have better effects both on the skin and as preservatives in the product [11].
Therefore, this study was developed to evaluate the antimicrobial and antioxidant activities of the essential oil from the fresh leaves of Siparuna guianensis. This species, known as ‘negramina’, occurs from Nicaragua to the north of South America and, in Brazil, is found mainly in the states of Mato Grosso do Sul, Mato Grosso, Goiás, Minas Gerais, Pará, Paraná, Distrito Federal, Pernambuco, and Rio de Janeiro [17,18,19]. Siparuna guianensis stands out in the field due to the strong aroma released when its leaves are crushed. Initially, the essential oil has an intense and unpleasant smell, with earthy, woody, and spicy notes. Over time, this intense smell persists, evolving into a characteristic herbaceous aroma. In folk medicine, it has been used in the treatment of stomach disorders, headaches, rheumatism, fevers, and hypertension, and shows anti-inflammatory action [19]. Additionally, the volatile nature of essential oils makes them excellent agents for use as fragrances in cosmetics. However, they can also show anti-aging, anti-acne, antimicrobial, skin lightening, and sunscreen effects; these activities can enhance the dermato-cosmetic properties of the final products making them highly valuable for the cosmetic sector [20].
Thus, in this work, the study of the chemical composition and antimicrobial and antioxidant activities of the essential oil from the fresh leaves of S. guianensis was carried out for possible application in cosmetic formulations. A product prototype formulated from the oil in synergy with commercial antibiotics was also investigated.

2. Materials and Methods

2.1. Materials

The reagents 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS), 2,2-diphenyl-1-picrylhidrazyl (DPPH), and potassium persulfate were purchased from Sigma-Aldrich Co. (St. Louis, MO, USA). The tert-butyl hydroxytoluene (BHT), 6-hydroxy-2,5,7,8 tetramethylchromane-2-carboxylic acid (TROLOX), and n-alkane standard solutions (C9–C22) were also provided from Sigma-Aldrich Co. (St. Louis, MO, USA), while the L-ascorbic acid standard was purchased from LabSynth (Diadema, SP, Brazil). The solvents UV–vis-grade ethanol and chloroform were obtained by Tedia Company (California City, CA, USA). The antibiotics and antibacterial culture media were purchased from Sigma Aldrich Co. (St. Louis, MO, USA). Clinical Staphylococcus aureus (resistant to clindamycin, erythromycin, and penicillin G), Staphylococcus epidermidis A (extensively drug-resistant, XDR, resistant to ciprofloxacin, clindamycin, erythromycin, gentamicin, linezolid, oxacillin, and trimethoprim/sulfamethoxazole) and B (multidrug resistant, MDR, resistant to ciprofloxacin, erythromycin, gentamicin, and oxacillin) were provided by the Clinical Analysis Center of the University Hospital of the Federal University of Mato Grosso do Sul (Campo Grande, MS, Brazil). Staphylococcus pseudointermedius (resistant to amoxicillin + clavulanic acid, gentamicin, neomycin, azithromycin, cephalexin, cephalothin, streptomycin, and marbofloxacin) was provided by the Faculty of Veterinary Medicine and Animal Science at the Federal University of Mato Grosso do Sul. The commercial vehicle used in the preparation of the ointments was purchased from the magistral pharmacy Dermativa®, under the name ‘base ointment’, free of any bioactive components.

2.2. Samples

Leaves of S. guianensis A. DC. (Siparunaceae) were collected in Campo Grande, MS, Midwest of Brazil, in December 2023, on the Universidade Federal de Mato Grosso do Sul (UFMS) campus from a previously marked individual, being an exsiccate specimen identified by Professor Flávio M. Alves (UFMS) and deposited under the code 74331 in the UFMS Herbarium (coordinates: 20°30′30.1″ S, 54°36′58.1″ W). The Brazilian license for research on Brazil’s biodiversity is #A1D8864.

2.3. Essential Oil Extraction

Fresh leaves of S. guianensis (9.16 Kg) were extracted for 4 h by steam drag using stainless steel distillation equipment (Linax Indústria e Comércio LTDA, São Paulo, Brazil), with a capacity of 20 kg of raw material, to yield 0.0201% w/w of essential oil. The method used to extract the essential oil was steam distillation. The light yellow essential oil was dried over anhydrous sodium sulfate (Na2SO4), packaged in amber glass bottles, and then stored in a freezer at −15 °C until analysis.

2.4. Analysis of the Oil

2.4.1. Gas Chromatography/Mass Spectrometry (GC/MS)

The essential oil was analyzed in a GC 2010 high-resolution gas chromatograph coupled to a QP-2010 Plus mass spectrometer (Shimadzu Corporation, Kyoto, Japan) equipped with an AOC-20i autosampler (Shimadzu Corporation, Kyoto, Japan). An Rtx-5MS (Restek®, Bellefonte, PA, USA) chromatographic column (5% diphenyl–95% dimethylpolysiloxane) of 30 m, 0.25 mm i.d., with a 0.25 mm film thickness, was utilized.

2.4.2. Chromatography Conditions

The essential oil was dissolved in dichloromethane (1 mg·mL−1) and a volume of 1 µL in splitless mode, at a ratio of 1:50, was injected. The injector and detector temperatures were 250 and 260 °C, respectively, with the carrier gas being He (White Martins®, analytical grade) at a flow rate of 1 mL·min−1, and a pressure of 86.7 kPa. The oven temperature was programmed from 60 °C to 320 °C at a rate of 4 °C/min to 320 °C and then kept isothermal for 20 min at 320 °C. The spectra were acquired using the electron impact technique at an impact energy of 70 eV, with mass range of 45 to 450 Da. The same chromatographic conditions were used to inject a mixture of linear hydrocarbons (C9 to C22 alkanes). The identification of the components in the essential oil was based on a comparison of the mass spectra obtained with those of commercial mass spectral libraries (Wiley 7 lib and Nist 08 lib) and also by using the Retention Index (RI) calculated for each constituent and comparing them with the RIs available in the literature [21]. The chromatograms and mass spectra were generated by GCMS Postrun Analysis software v. 2.72 (Shimadzu Corporation, Kyoto, Japan).

2.5. Ointment Preparation

S. guianensis essential oil (EO) was incorporated to a commercial vehicle prepared with BHT, disodium EDTA, cetearyl alcohol, sodium cetearyl sulfate, ceteareth 20, imidazolidinyl urea, mineral oil, petrolatum, and demineralized water. Some ointment formulations were prepared as follows, using ampicillin (AMP) and also gentamicin (GENTA) (the amounts of antibiotic and essential oil are described per gram of ointment): (1) 0.5 mg GENTA + 2.5 mg EO/g; (2) 1 mg GENTA + 2.5 mg EO/g; (3) 0.5 mg GENTA + 10 mg EO/g; (4) 1 mg GENTA + 10 mg EO/g; (5) 2 mg GENTA + 10 mg EO/g; (6) 1 mg GENTA + 20 mg EO/g; (7) 0.5 mg GENTA/g; (8) 1 mg GENTA/g; (9) 2 mg GENTA/g; (10) 2.5 mg EO/g; (11) 10 mg EO/g; (12) 20 mg EO/g. Finally, the samples were incorporated into the ointment until the complete homogenization of the ointment was achieved [22].

2.6. Antibacterial Properties of Essential Oil from the Leaves of S. guianensis

2.6.1. Determination of Minimal Inhibitory Concentration (MIC)

Antimicrobial activities of the individual essential oil and essential oil (EO) and the antibiotics ampicillin (AMP) and gentamicin (GENTA) were determined by broth microdilution method [23]. Two-fold dilutions were carried out in 96-well plates prepared with Mueller–Hinton broth (Sigma Aldrich) to produce a final concentration of 200 to 0.2 μg·mL−1 for commercial antibiotics and 4000 to 31.25 μg·mL−1 for the essential oil, obtaining a final volume of 100 μL in each well. Overnight cultures of each bacterial species on the Mueller–Hinton agar (Sigma-Aldrich) were used to prepare the bacterial inoculum. These cultures were then diluted in a saline solution (0.9%) to a concentration of approximately 108 CFU.mL−1 (0.5 on the McFarland scale). Then, each solution was diluted 1:10 in a saline solution and 5 µL was added to each well containing the test samples. For 18 hours, the microdilution trays were incubated at 36 °C. The experiments were performed in triplicate. Following this timeframe, each well received 20 μL of a 0.5% aqueous solution of triphenyl tetrazolium chloride (TTC), and the trays were incubated for two hours at 36 °C. Additionally, TTC changed from colorless to red in the wells where bacterial growth occurred. The minimal inhibitory concentration (MIC, µg.mL−1), was determined as the lowest concentration of the sample at which no color change was observed.

2.6.2. Synergy Testing

Using a standard checkerboard microtiter method [23], the synergism between essential oil and ampicillin (EO + AMP) and gentamicin (EO + GENTA) was investigated against S. aureus, S. pseudointermedius, and S. epidermidis (strains A and B). The essential oil was submitted to serial two-fold dilutions to obtain concentrations ranging from 4000 to 31.25 μg.mL−1 in Mueller–Hinton broth-prepared 96-well plates, with a final volume of 50 μL in each well. Next, 50 μL of the antibiotic solutions was added to each well, resulting in concentrations that ranged vertically between 200 and 0.2 μg.mL−1. The essential oil had final concentration values ranging from 2000 μg.mL−1 to 15.6 μg.mL−1. After preparing the bacterial inoculums as described earlier, 5 μL was applied to each well holding the test samples. The plates were then incubated at 36 °C for 18 h. Following the addition of TTC, the combinations’ MICs were determined. The culture medium was used as a negative control, DMSO was used as a blank, and the culture medium without inoculation was considered the sterility control.
The fractional inhibitory concentration (FIC) and the fractional inhibitory concentration index (FICI) were calculated using the following formulas:
FIC = Combined MIC of EO or antibiotic/Individual MIC of EO or antibiotic
FICI = FIC of EO + FIC of antibiotic
The FICI values were interpreted as follows: a synergic effect when FICI < 0.5, an additive effect when 0.5 < FICI ≤ 1, and indifferent effect at 1 < FICI ≤ 4, and an antagonist effect at FICI > 4 [24,25,26].

2.6.3. Ointment Testing

All prepared ointments were evaluated using an agar overlay assay [27], with the commercial base serving as the negative control. A sample of each ointment (approximately 60 mg) was transferred to a well, punched aseptically with a sterile tip (±8 mm) on a Mueller–Hinton agar plate. Subsequently, the plates were covered with soft Mueller–Hinton agar containing the tested bacteria (S. aureus and S. epidermidis B) and incubated at 36 °C for 18 h. After this period, inhibition zones were measured by eye using a ruler.

2.7. Evaluation of Antioxidant Activity of Essential Oil

2.7.1. DPPH Radical Scavenging Assay

The 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay was performed according to methods developed earlier [28,29]. The reaction was carried out with 500 μL of the sample at concentrations of 10.0–100 μg.mL−1 and 2.5 mL of a 0.1 mM DPPH solution. The solutions were prepared in ethanol. The samples were stored at room temperature in the dark for 30 min, followed by measuring the absorbance at 515 nm in a spectrophotometer (Kasvi®, with a UV–vis range of 190–1100 nm). Subsequently, the samples were kept in the dark at room temperature for 60 min, and after this period, the previous experiment was repeated. Ascorbic acid and TROLOX [(±)-6-hidroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid] served as positive controls and were analyzed under the same conditions. For the negative control, 2.5 mL of the 0.1 mM DPPH solution was mixed with 500 μL of ethanol. The experiments were conducted in triplicate. The inhibition of free radicals (DPPH), in percentage, by antioxidant (sample) and the amount of the sample (antioxidant) required to reduce the concentration of free radicals by 50% (IC50) were calculated according to Santos et al. (2023) [29].

2.7.2. ABTS Radical Scavenging Assay

The antioxidant activity of the essential oil of S. guianensis was measured by the ABTS+• radical cation decolorization assay [30,31,32]. The ABTS+• radical cation was prepared from a reaction of 5 mL of a 7 mM ABTS [2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt] stock solution with 88 μL of a 140 mM potassium persulfate solution. The mixture was kept in the dark, at room temperature, for 16 h. Then, 1 mL of this mixture was diluted in ethyl alcohol (ratio 1:88) until an absorbance of 0.70 ± 0.05 at 734 nm. Furthermore, 3.0 mL of this solution was added to glass tubes containing 300 μL of ethanol solutions of the essential oil at concentrations of 10 to 100 μg.mL−1, and the tubes were homogenized for 30 s. The samples were incubated in the dark and at room temperature for 6 min and 30 min; after the respective incubation periods, absorbance readings were taken at 734 nm. The control was formed by 3.0 mL of the ABTS radical cation solution and 300 µL of ethanol. All experiments were developed in triplicate. Tert-butyl hydroxytoluene (BHT) was used as a positive control. The percentage of inhibition (A%) of the ABTS cation radical by the antioxidant agent was calculated using the following equation
A% = [(Abscontrol − Abssample)/Abscontrol] × 100
where Abscontrol is the absorbance of the control solution and Abssample is the absorbance of the sample. The IC50 value was determined using linear regression of the graph between the percentage of ABTS cation radical scavenging and the concentration of the samples (µg.mL−1) [33].

3. Results and Discussion

3.1. Chemical Composition of Siparuna guianensis

In total, 30 chemical compounds were identified in the essential oil of S. guianensis, comprising 90.25% of the constituent volatiles present in the sample (Table 1). Sesquiterpene hydrocarbons were the predominant components of the essential oil (74.70%), presenting the major constituents bicyclogemacrene (32.52%), germacrene D (21.60%), and germacrene B (6.84%). We observed the presence of oxygenated sesquiterpenes (8.70%) such as α-muurolol (1.97%) and spathulenol (1.19%). Furthermore, only 5.09% of monoterpene hydrocarbons were detected, and myrcene was considered the majority compound with 3.66% (Table 1). The ion chromatogram in relation to the chemical composition is shown in Figure 1. The chemical composition of this oil differs from previous studies of the essential oil of leaves of the same species harvested in different biomes. Previous studies on plants from the northern region of Brazil, specifically the Amazon region, identified myrcene (79.47%), 2-undecanone (7.86%), germacrene D (3.12%), and germacrene B (1.17%) as the major constituents of S. guianensis leaf oil [34]. Another specimen collected in same biome from Belém (Pará, Brazil) showed atractylone (18.65%), trans-β-elemene (11.78%), germacrene D (7.61%), cruzerene (7.10%), γ-elemene (7.04%), and δ-elemene (5.38%) as the major components [35]. Martins et al. (2021) [36] identified iso-shiobunone (23.9%) and epi-shiobunone (18.9%) as the main components in a specimen collected in Cantá (Roraima State, Brazil), while Andrade et al. (2013) [37] reported, in a sample collected in Minas Gerais (Brazil) in the Atlantic Forest area, the presence of the monoterpene β-myrcene (13.14%) and of the sesquiterpenes germacrene-D (8.68%) and bicyclogermacrene (16.71%) as the main constituents. In all samples, spathulenol was identified in low concentrations [34,36,37], but it was absent in the sample collected in the state of Pará, Brazil [35]. On the other hand, compounds such as atractylone, cruzerene, iso-shiobunone, and epi-shiobunone, which were the main components of S. guianensis specimens collected in other regions of Brazil [35,37], are absent in this essential oil sample (Table 1). It is worth mentioning that this is the first study on S. guianensis from the Cerrado region.
Variations in the chemical composition of essential oils can be attributed to various factors, including the plant development stage, circadian rhythms, harvesting season, nutrient and water availability, altitude, temperature, adaptation mechanisms, location, UV radiation, the essential oil extraction method [38,39], geographical sources, genetics, soil characteristics, and environmental factors at the collection sites [39,40]. These factors may also account for the presence or absence of some constituents in the essential oil sample of this work compared with previous investigations of the chemical composition [34,35,36].

3.2. Antimicrobial Assays

The essential oil from the fresh leaves of S. guianensis was investigated for its antimicrobial activity against clinically resistant strains of S. aureus, S. pseudointermedius and S. epidermidis, alone and in combination with ampicillin and gentamicin, which are commercial antibiotics.
Staphylococcus aureus is a notable bacterium for its ability to cause purulent skin and soft tissue infections [41], as well as other infections [29]. Additionally, Staphylococcus pseudointermedius, which rarely causes infection in humans, is far more common in animals, such as dogs. However, the most frequent clinical presentations are skin abscesses and wound infections in humans [29,42,43]. Moreover, S. epidermidis is typically recognized as a commensal bacterium and is one of the most common and universally present bacterial species on human skin and mucous membranes. They can colonize medical implants such as cerebrospinal fluid shunts, intraocular lenses, and prosthetic joints, among others [29,44]. Thus, the antimicrobial activity of essential oil from fresh S. guianensis leaves was investigated against three bacteria known to cause skin infections.
According to Table 2, the essential oil of S. guianensis showed antimicrobial activity only against S. epidermidis strain A (MIC = 500 µg.mL−1). Moreover, the combination of essential oil (EO) and ampicillin (AMP), when tested against S. epidermidis strain A (XDR strain), resulted in a synergistic antibacterial effect (FICI = 0.375), reducing the isolated antibiotic MIC by four-fold (a change in MIC from 6.25 µg.mL−1 when alone to 1.56 µg.mL−1 when in combination with the EO). The most pronounced synergistic effect was demonstrated by essential oil and AMP when assayed against S. aureus, which showed a fractional inhibitory concentration index value of 0.256 and a reduction in the antibiotic MIC from 3.125 µg.mL−1 to 0.780 µg.mL−1 (Table 2). Additionally, when tested against S. epidermidis strain B (MDR strain) and S. pseudointermedius, the combination of EO and AMP resulted in an additive antibacterial effect (FICI = 0.503) (Table 2). The antimicrobial potential of the essential oil from leaves of S. guianensis may be attributed to the significant sesquiterpene hydrocarbon concentration (74.70%), among other terpenoids with the lowest quantity. Also, the co-occurrence of the major constituents bicyclogermacrene (32.52%) and germacrene D (21.60%) found in the essential oil from leaves of S. guianensis has been reported in essential oils with antibacterial properties [35]. Gentamicin (GENTA) was also chosen for this study, since it is already used in topical formulations, being stable in creams and ointments. However, only S. epidermidis strains were evaluated for combinations with gentamicin, as the other bacteria were sensitive to this drug. When in combination with GENTA, the S. guianensis essential oil acted synergistically with the antibiotic against S. epidermidis B, showing a four-fold decrease in the MIC values of the combined components (Table 2). For S. epidermidis A, the observed effect was additive.
Akgul et al. (2021) [45] showed that cosmetic products can be a source of bacteria with acquired resistance to antibiotics. They identified several bacteria in cosmetic products, including the multidrug-resistant bacteria S. aureus and S. epidermidis. Therefore, the results obtained by Akgül et al. (2021) [45] and those obtained in this work suggest that the combination of essential oil from S. guianensis and conventional antibiotics may be an effective solution to develop dermocosmetics with greater efficacy against multidrug-resistant bacteria, ensuring the safety and durability of products. Furthermore, natural products with antimicrobial properties can be useful, especially for the development of antiseptic skin products that inactivate, reduce, prevent, or stop the growth of microorganisms with the intention of mitigating or preventing diseases [1]. Thus, natural products can replace synthetic agents used to preserve cosmetics from pathogenic microorganisms, increasing consumer interest in products containing active agents of natural origin [13].
With this in mind, several ointments were prepared containing different amounts of S. guianensis EO, gentamicin, and/or a combination of both. The ointments were evaluated against S. aureus (susceptible to gentamicin) and S. epidermidis B (resistant to gentamicin) (Table 3). Ointments 7, 8, and 9, containing only gentamicin, showed activity against S. aureus, with inhibition zones of 15 mm, 19 mm, and 22 mm, respectively. For S. epidermidis B, only Ointment 9 was active, with a zone of 15 mm (2 mg GENTA/g). Commercial products based on gentamicin are marketed at a concentration of 1 mg of gentamicin/g. For Ointments 10, 11, and 12, with only EO, there was no activity (Figure S1, Supplementary Material).
For Ointments 1 to 6, containing different combinations of EO and GENTA, promising results were observed. For S. aureus, Ointment 3, with 0.5 mg GENTA + 10 mg EO, showed an inhibition zone of 20 mm, comparable with the zone observed for the ointment containing 2 mg GENTA (Ointment 9). In the case of S. epidermidis, the combination of EO with gentamicin showed some activity for the ointments with 1 mg of gentamicin and 20 mg of EO (Ointment 6), and for the combination of 2 mg of gentamicin and 10 mg of EO (Ointment 5), for which inhibition zones of 10 mm were observed. Ointments 10–12 formulated only with EO did not show activity in this assay. Therefore, it is interesting to highlight that, for some of the tested formulations, the combination of S. guianensis EO improved the activity of the antibiotic gentamicin, making it possible to reach the same effect on the bacteria at a lower concentration, while adding a pleasant herbal scent, making the product more commercially attractive. These results are noteworthy, given the intrinsic resistance of S. epidermidis to the antibiotic, indicating that the synergy between EO and gentamicin can overcome resistance barriers.
This approach not only improves the antibiotic’s efficacy against resistant strains but also provides a pathway to reducing antibiotic usage, a key driver of bacterial resistance. The incorporation of essential oil (EO) in the tested vehicle delivered a dual advantage: enhancing antimicrobial activity, thereby promoting the sustainable use of antibiotics and imparting a pleasant fragrance to the formulation. As a future perspective, the antibiofilm properties of this oil should be investigated, as such findings could broaden its potential applications in topical formulations.

3.3. Antioxidant Activity

The essential oil from the fresh leaves of S. guianensis was investigated as an anti-aging (antioxidant) agent for cosmetic purposes using DPPH radical scavenging and ABTS+• radical cation scavenging assays. This oil, rich in sesquiterpene hydrocarbons (78.36%), which showed, as the main constituents, bicyclogemacrene (32.52%), germacrene D (21.60%), and germacrene B (6.84%), was assessed via the DPPH technique, but no antioxidant activity was observed [46,47] (Table 4).
The DPPH technique is widely used to assess the free radical scavenging activity of antioxidants. This method relies on the reduction in color intensity, which occurs when antioxidant substances transfer an electron or donate a hydrogen radical to the DPPH molecule, neutralizing its unpaired electron [28,46,47,48,49,50,51]. According to Mata et al. (2007) [51], the fact that terpene compounds are incapable of donating a hydrogen atom explains why there was no antioxidant activity seen in the DPPH reduction. Additionally, the fact that the essential oil from the fresh leaves of S. guianensis does not show antioxidant activity can be explained by how this essential oil is composed of 83.45% monoterpene and sesquiterpene hydrocarbons. On the other hand, lipid systems that contain a low amount of monoterpene and sesquiterpene hydrocarbons, and a significant amount of the phenolic monoterpenes thymol and carvacrol may present antioxidant activity against the DPPH radical, since the last two monoterpenes act as antioxidants in several essential oils [52,53,54].
Antioxidant activity is a complex process that can take place via various pathways. Because of its complexity, assessing the antioxidant activity of individual compounds or mixtures of substances requires the execution of multiple tests [22]. Thus, the ABTS+• radical cation scavenging assay was the second test used. One benefit of the ABTS+• radical cation is that it can be used to determine both lipophilic and hydrophilic antioxidants [46]. In addition, both radicals have the ability to take electrons and radical hydrogen from the antioxidant compounds present in the samples. There is a close link between the suppression of ABTS+• radical cation activity in an antioxidant sample and the radical scavenging capacity of DPPH [54]. The antioxidant activity of the examined essential oil was also not detected in the concentration ranges tested using the ABTS method. The sample showed an IC50 value greater than 500 μg.mL−1, while the IC50 value of the tert-butyl hydroxytoluene (BHT) standard was 29.38 μg.mL−1 (Table 3). Therefore, the absence of antioxidant activity in S. guianensis essential oil can also be justified by the majority presence of hydrocarbon terpenes in its composition (Table 1), which do not donate radical hydrogen to the ABTS radical cation [55,56].

4. Conclusions

Thirty compounds were identified in the essential oil extracted from the leaves of Siparuna guianensis. The major classes of compounds were sesquiterpene hydrocarbons (74.79%), followed by oxygenated sesquiterpenes (8.70%) and monoterpenes (5.09%). The antioxidant activity of the essential oil was assessed using DPPH and ABTS radical scavenging assays; however, no significant antioxidant capacity was observed in either test. Furthermore, this study represents the first report on the antimicrobial activity of S. guianensis essential oil against Staphylococcus epidermidis and S. pseudointermedius, both of which are associated with skin infections. The essential oil exhibited a significant synergistic effect against S. aureus and S. epidermidis (Strain A, XDR) when combined with ampicillin, and a similar effect against S. epidermidis (strain B, MDR) when combined with gentamicin, suggesting that this species is a rich source of compounds with antimicrobial potential. Additionally, the formulation of ointments containing the essential oil and gentamicin yielded promising results, enhancing antimicrobial activity compared with gentamicin alone. Notably, the combination EO–gentamicin reduced the required antibiotic concentration by half compared with commercial ointments while also restoring gentamicin’s efficacy against multidrug-resistant bacteria, for which the antibiotic was previously ineffective. These findings underscore the potential of S. guianensis as a valuable natural resource for the development of novel therapeutic agents and dermocosmetic formulations. In this context, further studies on the in vivo efficacy, transdermal action, and toxicity are necessary to advance the development of these formulations.

Supplementary Materials

The following supporting information can be downloaded at https://www.mdpi.com/article/10.3390/cosmetics12020081/s1. Figure S1. Plates and zones of inhibition for ointment formulations with Siparuna guianensis leaf EO and gentamicin (GENTA) against a clinical sensitive strain of S. aureus (SAC) and MDR S. epidermidis (176H): (1) 0.5 mg GENTA + 2.5 mg EO/g; (2) 1 mg GENTA + 2.5 mg EO/g; (3) 0.5 mg GENTA + 10 mg EO/g; (4) 1 mg GENTA + 10 mg EO/g; (5) 2 mg GENTA + 10 mg EO/g; (6) 1 mg GENTA + 20 mg EO/g; (7) 0.5 mg GENTA/g; (8) 1 mg GENTA/g; (9) 2 mg GENTA/g; (10) 2.5 mg EO/g; (11) 10 mg EO/g; (12) 20 mg EO/g.

Author Contributions

Conceptualization, N.C.Y. and É.L.d.S.; methodology, A.C.M., G.d.S.d.J. and J.M.F.S.; data curation, É.L.d.S. and A.C.M.; writing—original draft, É.L.d.S.; writing—review and editing, A.C.M., É.L.d.S., and N.C.Y.; project administration, N.C.Y. and É.L.d.S.; supervision, N.C.Y. All authors have read and agreed to the published version of the manuscript.

Funding

The Fundação de Apoio ao Desenvolvimento do Ensino, Ciência e Tecnologia do Estado de Mato Grosso do Sul—FUNDECT (Project No. 342/2022 to process No. 71/038.838/2022; Project No. 308/2022 to process No. 71/032.451/2022) provided financial assistance for this study. The Coordenação de Aperfeiçoamento de Pessoal de Nível Superior—Brasil (CAPES) (Finance Code 001) provided partial funding for this work.

Data Availability Statement

Dataset available on request from the authors.

Acknowledgments

The authors are grateful to the Fundação Universidade Federal de Mato Grosso do Sul, FUNDECT-MS, CNPq, and CAPES for the financial support.

Conflicts of Interest

The authors declare no conflicts of interest.

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Cosmetics 12 00081 g001
Figure 1. Total ion chromatogram of essential oil from the fresh leaves of S. guianensis. The numbers refer to the compounds identified in Table 1.
Figure 1. Total ion chromatogram of essential oil from the fresh leaves of S. guianensis. The numbers refer to the compounds identified in Table 1.
Cosmetics 12 00081 g001
Table 1. Chemical composition of the essential oil from fresh leaves of S. guianensis.
Table 1. Chemical composition of the essential oil from fresh leaves of S. guianensis.
PeakRt (min)CompoundsMolecular
Formula		RIlitRIexpPeak Area (%)
15.531β-PineneC10H169749690.11
25.794MyrceneC10H169889793.66
36.780LimoneneC10H16102410261.16
47.283E-OcimeneC10H16104410440.16
514.8742-UndecanonaC11H22O129312931.76
616.286δ-ElemeneC15H24133513382.30
717.523α-CopaeneC15H24137413770.66
817.810β-BourboneneC15H24138713860.40
918.027β-ElemeneC15H24138913931.88
1018.890E-CaryophylleneC15H24141714211.31
1119.191β-GurjuneneC15H24143114310.19
1219.312γ-ElemeneC15H24143414352.11
1319.495AromadendreneC15H24143914410.25
1419.939α-HumuleneC15H24145214550.44
1520.167Allo-aromadendreneC15H24145814620.28
1620.560Cadina-1(6),4-dieneC15H24146114760.17
1720.652γ-MuuroleneC15H24147814790.29
1820.802Germacrene DC15H241484148421.60
1920.948β-SelineneC15H24148914880.48
2021.292BicyclogermacreneC15H241500150032.52
2121.404(Z)-α-bisaboleneC15H24150615041.11
2222.049δ-CadineneC15H24152215261.87
2323.052Germacrene BC15H24155915606.84
2423.649SpathulenolC15H24O157715801.19
2523.833GlobulolC15H26O159015870.80
2624.070ViridiflorolC15H26O159215950.66
2725.379IsosphatulenolC15H24O163316420.99
2825.482τ-MuurololC15H26O164016450.95
2925.597α-MuurololC15H26O164416491.97
3025.830α-CadinolC15H26O165216582.14
Total identified (%)90.25
Monoterpene hydrocarbons (%)5.09
Sesquiterpene hydrocarbons (%)74.70
Oxygenated sesquiterpenes (%)8.70
Others (%)1.76
RIlit, retention index from the literature; RIexp, experimental retention index (calculated). Components ≤ 0.1% were not listed.
Table 2. Determination of the minimum inhibitory concentration (MIC, in µg.mL−1) of antibiotics and essential oil from the fresh leaves of Siparuna guianensis, the fractional inhibitory concentration (FIC), and the fractional inhibitory concentration index (FICI) of antibiotics and essential oil combined against drug-resistant bacteria.
Table 2. Determination of the minimum inhibitory concentration (MIC, in µg.mL−1) of antibiotics and essential oil from the fresh leaves of Siparuna guianensis, the fractional inhibitory concentration (FIC), and the fractional inhibitory concentration index (FICI) of antibiotics and essential oil combined against drug-resistant bacteria.
PathogensSample/CombinationIndividual MICCombined MICFICFICIEffect
Staphylococcus
epidermidis (A)		EO50062.50.125
AMP6.251.560.25 Synergism
EO + AMP 0.375
EO50031.250.06
GENTA100500.5 Additivity
EO + GENTA 0.506
Staphylococcus
epidermidis (B)		EO100031.250.125
AMP3.1251.560.25 Additivity
EO + AMP-- 0.503
EO10002500.25
GENTA6.251.560.25 Synergism
EO + GENTA 0.5
Staphylococcus aureusEO≥100062.50.06
AMP3.1250.782.5 Synergism
EO + AMP 0.256
Staphylococcus
pseudointermedius		EO≥100031,250.03
AMP50250.5 Additivity
EO + AMP 0.503
AMP, ampicillin; GENTA, gentamicin; EO, essential oil from S. guianensis. Synergic effect (FICI < 0.5); additive effect (FICI > 0.5–1); indifferent effect (FICI > 1–4); antagonist effect (FICI > 4).
Table 3. Zones of inhibition for ointment formulations with Siparuna guianensis leaf EO and gentamicin (GENTA) against a clinical sensitive strain of S. aureus and MDR S. epidermidis.
Table 3. Zones of inhibition for ointment formulations with Siparuna guianensis leaf EO and gentamicin (GENTA) against a clinical sensitive strain of S. aureus and MDR S. epidermidis.
SampleOintment
Formulation		Antimicrobial Activity
(Inhibition Zones)
S. aureusS. epidermidis
10.5 mg GENTA + 2.5 mg EO/g14 mmN.O.
21 mg GENTA + 2.5 mg EO/g18 mmN.O.
30.5 mg GENTA + 10 mg EO/g20 mmN.O.
41 mg GENTA + 10 mg EO/gN.O.N.O.
52 mg GENTA + 10 mg EO/g19 mm10 mm
61 mg GENTA + 20 mg EO/g22 mm10 mm
70.5 mg GENTA/g15 mmN.O.
81 mg GENTA/g19 mmN.O.
92 mg GENTA/g22 mm15 mm
N.O., not observed.
Table 4. Determination of antioxidant activity of essential oil from Siparuna guianensis leaves.
Table 4. Determination of antioxidant activity of essential oil from Siparuna guianensis leaves.
SampleAntioxidant Activities (IC50, µg.mL−1)
DPPHABTS
EO>500>500
BHT a-29.38
Ascorbic acid a37.15-
TROLOX a27.02-
EO, essential oil from Siparuna guianensis; IC, inhibitory concentration; DPPH, 2,2-diphenyl-1-picrylhydrazyl; ABTS, 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic) acid; BHT, tert-butyl hydroxytoluene; TROLOX, (6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid). (-): not tested. Positive controls a.
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dos Santos, É.L.; Siqueira, J.M.F.; da Silva de Jesus, G.; Micheletti, A.C.; Yoshida, N.C. Potential of Essential Oil from Siparuna guianensis A. DC. (Siparunaceae) as an Antimicrobial Adjuvant in Topical Formulations. Cosmetics 2025, 12, 81. https://doi.org/10.3390/cosmetics12020081

AMA Style

dos Santos ÉL, Siqueira JMF, da Silva de Jesus G, Micheletti AC, Yoshida NC. Potential of Essential Oil from Siparuna guianensis A. DC. (Siparunaceae) as an Antimicrobial Adjuvant in Topical Formulations. Cosmetics. 2025; 12(2):81. https://doi.org/10.3390/cosmetics12020081

Chicago/Turabian Style

dos Santos, Érica Luiz, Juliana Mendes Franco Siqueira, Genilson da Silva de Jesus, Ana Camila Micheletti, and Nídia Cristiane Yoshida. 2025. "Potential of Essential Oil from Siparuna guianensis A. DC. (Siparunaceae) as an Antimicrobial Adjuvant in Topical Formulations" Cosmetics 12, no. 2: 81. https://doi.org/10.3390/cosmetics12020081

APA Style

dos Santos, É. L., Siqueira, J. M. F., da Silva de Jesus, G., Micheletti, A. C., & Yoshida, N. C. (2025). Potential of Essential Oil from Siparuna guianensis A. DC. (Siparunaceae) as an Antimicrobial Adjuvant in Topical Formulations. Cosmetics, 12(2), 81. https://doi.org/10.3390/cosmetics12020081

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