Next Article in Journal
Pseudomonas cannabina pv. alisalensis Virulence Factors Are Involved in Resistance to Plant-Derived Antimicrobials during Infection
Next Article in Special Issue
Effect of Tecoma stans (L.) Juss. ex Kunth in a Murine Model of Metabolic Syndrome
Previous Article in Journal
Genomic Prediction Accuracy of Stripe Rust in Six Spring Wheat Populations by Modeling Genotype by Environment Interaction
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Plants
Volume 11
Issue 13
10.3390/plants11131739
plants-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Communication

Eupatorin and Salviandulin-A, with Antimicrobial and Anti-Inflammatory Effects from Salvia lavanduloides Kunth Leaves

by
Manasés González-Cortazar
1,
David Osvaldo Salinas-Sánchez
2,3,*,
Maribel Herrera-Ruiz
1,
Dionisio Celerino Román-Ramos
3,
Alejandro Zamilpa
1,
Enrique Jiménez-Ferrer
1,
Ever A. Ble-González
4,
Patricia Álvarez-Fitz
5,
Ricardo Castrejón-Salgado
6 and
Ma. Dolores Pérez-García
1,*
1
Centro de Investigación Biomédica del Sur, Instituto Mexicano del Seguro Social, Argentina No. 1, Col. Centro, Xochitepec 62790, Morelos, Mexico
2
Centro de Investigación en Biodiversidad y Conservación (CIByC), Universidad Autónoma del Estado de Morelos (UAEM), Av. Universidad 1001, Col. Chamilpa, Cuernavaca 62209, Morelos, Mexico
3
Escuela de Estudios Superiores del Jicarero (EESJ), Universidad Autónoma del Estado de Morelos, Carretera Galeana-Tequesquitengo s/n Col. el Jicarero, Jojutla 62909, Morelos, Mexico
4
División Académica de Ciencias Básicas, Universidad Juárez Autónoma de Tabasco, Carretera Cunduacán-Jalpa Km. 0.5, Cunduacán 86690, Tabasco, Mexico
5
Laboratorio de Toxicologia, Cátedra CONACyT-Universidad Autónoma de Guerrero, Av. Lázaro Cárdenas s/n. Col. La Haciendita, Chilpancingo 39070, Guerrero, Mexico
6
Instituto Mexicano del Seguro Social, Unidad de Medicina Familiar Número 3, Avenida Insurgentes Esquina con Emiliano Zapata s/n., Centro, Jiutepec 62550, Morelos, Mexico
*
Authors to whom correspondence should be addressed.
Plants 2022, 11(13), 1739; https://doi.org/10.3390/plants11131739
Submission received: 31 May 2022 / Revised: 23 June 2022 / Accepted: 26 June 2022 / Published: 30 June 2022
(This article belongs to the Special Issue Biological and Chemical Activity of Metabolites of Medicinal Plants)
Browse Figures
Review Reports Versions Notes

Abstract

:
This study describes the antimicrobial and anti-inflammatory effects from extracts obtained from the leaves of Salvia lavanduloides. The plant material was macerated with three solvents of ascending polarity (n-hexane (Sl-Hex), ethyl acetate (Sl-AcOEt), and dichloromethane (Sl-D)). The extracts, fractions (SlD-2 and SlD-3), and isolated compounds (15,16-epoxy-10-β-hydroxy-neo-cleroda-3,7,13(16),14-tetraene-17,12R:18,19-diolide (1), salviandulin A (2), and eupatorin (3)) were evaluated as antimicrobials against Gram-negative, Gram-positive bacteria and the fungus Candida albicans (Ca) using the minimum inhibitory concentration (MIC) and the anti-inflammatory activity induced by 13-acetate of 12-O-tetradecanoylforbol (TPA). Sl-D and Sl-AcOEt extracts, SlD-2 and SlD-3 fractions showed the highest antimicrobial activity. The isolated compounds showed good activity against Pseudomonas aeruginosa with a MIC < 2 μg/mL, while the anti-inflammatory activity, the Sl-Hex, Sl-D extracts, and SlD-3 fraction presented an inhibition of 62, 45 and 61%, respectively, while (2) 70% and (3) 72%.

1. Introduction

One of the largest problems faced by the public health sector is infections transmitted within hospitals, Staphylococcus aureus being the main cause of infections after surgery, followed by pneumonia and intrahospital bacteremia, causing up to 50% mortality [1]. In Mexico, healthcare-related infections caused 32 deaths per 100,000 inhabitants, and it is estimated that prevalence can reach up to 23% in intensive care units [2]. It is generally known that many microorganisms are achieving antibiotic resistance, including Staphylococcus aureus and Pseudomonas aeruginosa. Therefore, the WHO has created a list of pathogens that need new pharmaceutical drugs to mitigate this problem [3]. S. aureus is an opportunistic pathogen, and it is one of the most dangerous staphylococcal bacteria. This Gram-positive bacterium can be part of the human microflora; it has been found on the skin and nasopharynx, affecting immunocompromised patients, causing skin, nose, urethra, vagina, and gastrointestinal tract infections. If mistreated, those infections can lead to pneumonias, endocarditis, and bone infections [4]. P. aeruginosa is also an opportunistic bacterium, easily infecting vulnerable patients with severe medical problems, such as otitis externa, ecthyma gangrenous, folliculitis, soft tissue infection, among others [5]. The moment a tissue is affected by any pathogenic bacteria, the response of cytokines, chemokines, lipid mediators, and bioactive amines is activated by various types of cells, such as monocytes, dendritic cells, natural killer cells, and mast cells, activating the release of pro-inflammatory molecules [6]. Inflammation is a process involved in the pathogenesis of bacterial infections, and approximately 20% of cancer-inducing inflammation cases have microbial organisms as the causative agent [7]. The antibiotics used to treat diseases caused by bacteria are β-lactams, such as penicillins, cephalosporins, monobactams, carbapenems, glycopeptides, aminoglycosides, and quinolones [8]. The use of NSAIDs (diclofenac, indomethacin) is a staple in medicine, although overuse can lead to undesirable side effects [9]. There is a variety of medicinal plants in Mexico that are used for the treatment of these conditions [10,11].
A wide variety of medicinal applications have been attributed to species of the genus Salvia. This genus is the most diverse of the family Lamiaceae, with approximately 1000 species distributed in different areas of the planet [12]. Central and South America alone have 500 species. Salvia is the second most diverse genus in Mexico, where 300 species are endemic to the country [13,14]. Its greatest distribution occurs in mountainous areas [15,16], such as the Occidental Sierra Madre, Eastern Sierra Madre, and the highlands of Chiapas [17]. It has been reported that some species of this genus possess properties such as antimicrobial, spasmolytic, central nervous system depressants, antitumor, and anti-inflammatory [18,19,20].
Salvia lavanduloides Kunth, an endemic plant of Mexico, is distributed around 21 states of the Republic, from Sinaloa and Durango to Oaxaca, Veracruz, and Chiapas. In Mexico, S. lavanduloides has several common names, such as lucema, mazorquilla, alucema, aguanta-tsitsiki, maroon chia, flor de cielo, and k’uironi simarroni [21]. This plant is used in Mexican traditional medicine to wash wounds and as an antipyretic. It has also been used to help with gastrointestinal issues, such as stomach and gallbladder ailments, toothaches, diarrhea, and vomiting. The leaves have also been used to treat gynecological conditions, such as menstruation, hemorrhaging, and abnormal vaginal discharges [22,23,24,25,26]. As ethnomedical use indicates, some of the problems for which the species is used are associated with bacterial infections and inflammation.
From this plant species, salviandulins A, B, and C (a group of secoclerodane-type diterpenoids) have been isolated and characterized [27,28]; however, there is only one pharmacological report evaluating eupatorin in the model of phorbel ester-induced pinna edema (TPA) [29]. The objective of the present work was to evaluate the antimicrobial and anti-inflammatory effect of extracts, fractions, and isolated compounds from the leaves of Salvia lavanduloides using the MIC method and phorbol ester-induced edema (TPA).

2. Results

2.1. Structural Elucidation of Compound (1), Salviandulin A (2), and Eupatorin (3)

Compound 1 (10 mg) was isolated as a white powder, soluble in methanol. The 13C NMR spectrum showed twenty signals, and DEPT analysis presented eight quaternary, six methines, five methylenes, and one methyl, indicating a diterpene. Based on spectroscopic data analysis (Table 1, see Supplementary Figures S1–S6) and the comparison with data described in the literature [30] this compound corresponds to a diterpene of type neo-clerodane known as 15,16-epoxy-10-β-hydroxy-neo-cleroda-3,7,13(16), 14-tetraene-17,12R:18,19-diolide (1). Figure 1.
Compound 2 (73.9 mg) was isolated as a white powder, and it is soluble in methanol. The 13C NMR spectrum showed twenty signals and two additional ones from an acetate group (CH3COO). According to the DEPT experiment, seven are quaternary, eight methines, four methylenes, and one methyl. Based on spectroscopic data analysis (Table 1, see Supplementary Data Figures S7–S12) and the comparison with data described in the literature [27], this compound corresponds to a diterpene known as salviandulin A (2). Figure 1.
Compound (3) was isolated as a yellow powder. 1H NMR spectra showed three ring systems: system A [δ 6.86 (1H, s, H-8)]; aromatic ABX system [δ 7.48 (1H, d, 1.4 Hz, H-2′), 7.55 (1H, dd, 1.4, 8.4 Hz, H-5′) and 7.10 (1H, d, 8.4 Hz, H-6′)]; and a double bond system in δ 6.86 (1H, s, H-3), indicating a flavone nucleus called 6-hydroxyluteolin. Additionally, three proton signals of oxygenated base were observed in δ 3.79, 3.92, and 3.97, corresponding to methoxy groups. Based on this information (see Supplementary Data, Figures S13–S18) the natural product was identified as 3′,5-dihydroxy-4′,6,7-trimethoxyflavone, or eupatorin (3), and was also consistent with spectroscopic data (see Table 2) described in the literature [31]. Figure 1.

2.2. Antimicrobial Activity of Extract, Fractions, and Compounds of S. lavanduloides

Table 3 shows the observed antimicrobial activity from the extracts against the strains evaluated. It is worth noting that, while Sl-AcOEt and Sl-D both showed antimicrobial activity against all strains tested, Sl-D presented the best activity with an MIC for Sa, SaR, Pa at 100, 100, and <25 μg/mL, respectively.
Table 4 shows the antibacterial activity shown by the fractions and compounds against the strains evaluated. The antimicrobial activity of the SlD-2 fraction shows that the inhibition of Sa, Se-2, Se-3, and Sh occurred at a concentration of 100 µg/mL, while for SaR and Se-1, it was 50 µg/mL. The antimicrobial activity in Ef, Pa, and Sd occurred at MIC < 25 µg/mL; however, in the case of Ca, the inhibition occurred at >200 µg/mL. For the SlD-3 fraction, the inhibition in Sa, SaR, Se-2, Se-3 was 200 µg/mL. For Sd and Ca, they presented an inhibition >200 µg/mL, highlighting that the MIC in Se-1, Ef, and Pa was <25 µg/mL. It should be noted that the antimicrobial activity of the SlD-2 and SlD-3 fractions in Ef and Pa was maintained with a MIC < 25 µg/mL, as in the Sl-D extract. However, when the isolated compounds were evaluated, the antimicrobial activity increased, obtaining those compounds 1–3 presenting an inhibition of 16 µg/mL for Sa, SaR, Se-1, Se-2, Se-3, Sh, Sd, only in compounds 2, 3 in Sd and Ca. It should be noted that the MIC < 2 µg/mL was obtained in Ef and Pa for the tree compounds, while in Sd, this concentration was obtained in compounds 1–3. The MIC < 2 µg/mL was in Ca when evaluating compound 1. The evaluation of the isolated compounds maintained the antimicrobial activity of the Sl-D extract.

2.3. Anti-Inflammatory Activity of Extracts, Fractions, and Compounds of S. lavanduloides

The local application of TPA in the mouse’s ear caused an edema of 11.76 mg, followed by administration of the non-steroidal anti-inflammatory drug (NSAID) indomethacin (INDO) to counteract it. Edema weight was reduced to 2.72 mg, representing an inhibition percentage of 76.87%. Table 5 presents the data of the anti-inflammatory activity of extracts, fractions, and compounds obtained from S. lavanduloides. The least polar extract (Sl-Hex) exhibited the highest anti-inflammatory activity, while Sl-D and Sl-AcOEt had a similar level of effect. The chemical separation was performer based on the results obtained from the antimicrobial activity; chemical separation of the dichloromethane extract was conducted. Two SlD-2 and SlD-3 fractions were generated, of which SlD-3 showed an anti-inflammatory effect. Continuing with the chemical separation of the most active fraction, SlD-3, the isolation of compounds 1, 2, and 3 was achieved. Mouse pinna edema was decreased with compounds 2 and 3, both showing similar activity when compared to the positive control indomethacin (Table 5).

3. Discussion

The interest in the phytochemical and pharmacological study of species of the genus Salvia has been provoked by its wide range of traditional medicinal uses in Mexico [32]. However, there have been no previous pharmacological references demonstrating the antimicrobial and anti-inflammatory effect of S. lavanduloides. In this work, it was demonstrated that all evaluated organic extracts of S. lavanduloides inhibited TPA-induced mouse ear edema. Dichloromethane extract (Sl-D) showed the highest antimicrobial activity, as well as significant anti-inflammatory activity. The phytochemical analysis of this extract allowed the isolation of the diterpene secoclerodane salviandulin A (2) and the flavone eupatorin (3). It is important to mention that both metabolites were previously identified in the genus Salvia [27,29,33].
In the TPA test, it was discovered that eupatorin (3) was the most active compound, followed by salviandulin A (2). This drug is widely used as a reference standard. The mechanism by which it exerts its anti-inflammatory effect is by inhibiting the enzyme cyclooxygenase (COX-1) [30]. The use of NSAIDs is a staple in medicine, although overuse can lead to undesirable side effects, such as gastric ulcers. Therefore, much of the medicinal plant research focuses on finding new anti-inflammatory treatments. For example, the gender Salvia has been used to treat inflammation, but only few pharmacological studies have been reported with this species [18,19,20].
These results are consistent with those previously reported [34]. Our experiment used the same model cited in the literature, except with a concentration of 1 mg/ear [29]. Comparing the percentages of inhibition of anti-inflammatory activity obtained, eupatorin (3) performed better in our experiment, with 72.45% inhibition of atrial edema versus 56.40% inhibition previously reported [29].
In recent decades, there has been an increase in the study of biological activities of diterpenes and flavones, such as salviandulin A (2) and eupatorin (3), which have been shown to have anti-inflammatory properties. In the case of eupatorin (3), inhibition of iNOS, COX-2, and tumor necrosis factor alpha (TNF-α) has been reported; in addition, it can also produce NO. It also inhibited carrageenan-induced swelling of mouse leg edema [35]. In the case of salviandulin A (2), none of its biological activities have been reported. Therefore, we are for the first time reporting that salviandulin A (2) exhibits anti-inflammatory properties. However, additional studies aimed at understanding the mechanisms of action of both salviandulin A (2) and eupatorin (3) are needed. In the literature, the isolation and structural elucidation of diterpenoid-type compounds, such as salviandulin E and D [27,28,31], have been reported; however, the metabolite reported here as salviandulin A has not been reported to date, and with it, no pharmacological activity. The findings documented herein support the use of S. lavanduloides in traditional medicine due to the isolated diterpenes and flavonoids presented in this plant, since antimicrobial and anti-inflammatory activity was proven.

4. Materials and Methods

4.1. Equipment and Reagents

NMR spectra were recorded on an Agilent DD2-600 at 600 MHz for 1H and 150 MHz for 13C NMR, using CDCl3 (compounds 1 and 2), CD3COCD3 (compound 3) as the solvent. Bidimensional experiments (COSY, HSQC, and HMBC). Chemical shifts are reported in ppm relative to TMS.

4.2. Plant Material

Salvia lavanduloides Kunth (10 plants, 1–1.5 m tall) leaves (all growth stages) were collected in San Juan Tlacotenco, Tepoztlán Morelos, in February 2020, and a specimen was deposited in the herbarium of the Center for Research in Biodiversity and Conservation (CIByC) of the Autonomous University of the State of Morelos (HUMO) with registration number: (HUMO-39806).

4.3. Extracts

The fresh leaves of S. lavanduloides (1 Kg) were dried at room temperature and stored away from light. The dry material (100 g) was crushed and macerated for 24 h sequentially (in triplicate) with two liters of ascending polarity solvents, i.e., n-hexane, ethyl acetate, and dichloromethane (Merck, Darmstadt, Germany). Each extract was filtered and concentrated under reduced pressure using a rota-evaporator (Heidolph at 50 °C) to obtain extracts Sl-Hex, Sl-AcOEt, and Sl-D, respectively.

4.4. Isolation and Identification of Compounds (1–3)

Sl-D extract (4.3 g) was adsorbed into 5 g of silica gel (silica gel 60, Merck, Darmstadt, Germany) and added to a glass column packed with silica gel (30 g, 70–230 mesh, Merck, Darmstadt, Germany). The circumvention system consisted of a gradient of hexane-ethyl acetate with polarity increments of 10%, collecting 36 fractions of 100 mL each. The fractions were concentrated at reduced pressure and monitored with normal phase chromatography and revealed with cerium sulfate. Fractions showing similarity in their chemical characteristics were grouped into 4 fractions (SlD-1 to SlD-4).
The SlD-3 fraction (584 mg) was then adsorbed in silica gel (RP-18, Merck, Darmstadt, Germany) for separation using a pre-packed reverse phase column (10 g, RP-18, Merck, Darmstadt, Germany) as a stationary phase. The elution system consisted of 100% acetonitrile (HPLC grade, Merck, Darmstadt, Germany) and polarity increments of 10% water, collecting volumes of 7 mL. Separation was monitored with reverse phase thin layer chromatography (RP-18, Merck, Darmstadt, Germany) and revealed with cerium sulfate. In fraction 21–22, the diterpene 15,16-epoxy-10-β-hydroxy-neo-cleroda-3,7,13(16), 14-tetraene-17,12R:18,19-diolide (1, 10 mg) was isolated. Fractions 26–28 had salviandulin A (2, 25 mg), and in fractions 37 and 38, a flavone called eupatorin (3, 85.2 mg) was identified (Figure 2).

4.5. Antimicrobial Activity

4.5.1. Bacterial

Extracts, fractions, and compounds were evaluated against Gram-positive bacteria: Staphylococcus aureus ATCC 29213 (Sa) and methicillin-resistant Staphylococcus aureus ATCC 42310 (SaR), Staphylococcus epidermis ATCC 35984 (Se-1), Staphylococcus epidermis ATCC 12228 (Se-2), Staphylococcus epidermis ATCC 1042 (Se-3), Staphylococcus haemolitycus ATCC 1165 (Sh), Enterococcus faecalis ATCC 29212 (Ef); and Gram-negative bacteria Pseudomonas aeruginosa ATCC 27853 (Pa), Salmonella dublin NTCC 9676 (Sd). In addition, all samples were also evaluated against the fungus Candida albicans ATCC 10231 (Ca).

4.5.2. Minimal Inhibitory Concentration (MIC)

Following a methodology previously described [36], all bacteria were standardized to Mc Farland’s 0.5 (1.5 × 108 mL/mL CFUs). The samples were diluted with dimethyl sulfoxide (DMSO, 200 μL), water (800 μL), and evaluated at concentrations of 200, 100, 50, and 25 μg/mL for extract and fractions, and pure compounds 16, 8, 4, and 2 μg/mL in microplates of 96 flat-bottomed wells, where 100 μL of Mueller Hinton broth (MHB, Merck, Darmstadt, Germany) and serial dilutions of the extracts were placed. An amount of 2 μL of bacterial strains were inoculated and incubated at 37 °C for 24 h. Gentamicin (10 μg/mL) was used as a positive control and DMSO as the negative control. All experiments were conducted in triplicate [36].

4.6. Anti-Inflammatory Activity

4.6.1. Animals

Male Balb-C mice (20 g) were obtained from Centro Médico Nacional SXXI-IMSS. They were organized in groups of 7 animals under controlled conditions with a 12 h light–dark cycle, at 20 ± 1 °C of temperature and free access to food (Labdiet 5008, Brentwood, MO, USA) and water. Mice were handled according to the Mexican Official Standard on the Care and Management of Animals (NOM-062-ZOO-1999) and international regulations. The Research Committee of IMSS approved the experimental designs used in the present study F-2021-1702-022.

4.6.2. 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced Mouse Ear Edema

Animal inflammation was induced following a method previously described [37,38]. Eight groups of seven mice each were formed. Then, TPA (Sigma Chemical Co. St. Louis, MO, USA, 2.5 g) dissolved in acetone (Baker, 20 L) was applied in the internal and external surface on the right ear to cause edema. Doses of 1 mg/ear of each treatment (Sl-Hex, Sl-AcOEt, Sl-D, SlD-2, SlD-3, salviandulin-A, eupatorin) were applied on the ear of each individual. Reference anti-inflammatory drug (indomethacin, Sigma Aldrich, St. Louis, MI, USA) was administered at 1 mg/ear. All treatments were dissolved in acetone and applied topically on ears immediately after the administration of TPA. Six hours after administration of the inflammatory agent, the animals were sacrificed by cervical dislocation. Circular sections of 6 mm in diameter were taken from both the treated (t) and the non-treated (nt) ears, which were weighed to determine the inflammation. The percentage of inhibition was obtained using the expression below:
Inhibition % = [(Δw control − Δw treatment/ΔW)] × 100
where Δw = wt − wnt; wt is the weight of the section of the treated ear; wnt is the weight of the section of the non-treated ear.

4.7. Statistical Analysis

Statistical evaluation was performed with SPSS (Version 22.0) software based on one-way ANOVA with a confidence level of 95%, * p ≤ 0.05, and followed by Dunnett’s test with one-tale: * p ≤0.05 in comparison with the negative control.

5. Conclusions

In this study, the anti-inflammatory and antimicrobial activity of the species Salvia lavanduloides Kunth was demonstrated by testing the three isolated compounds (diterpenoid (1), salviandulin A (2), and eupatorin (3)). Our data revealed that compound 1 was the most bioactive against all the tested microorganisms. It was also found that both salviandulin A and eupatorin present high anti-inflammatory activity at 70% and 72%, respectively, for mouse pinna edema. Furthermore, the elucidation of the chemical structures of the three isolated and characterized natural products (1 to 3) highlights the utility of the protocol utilized and opens up an opportunity to foster drug discovery, while validating the use of this plant in traditional medicine.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/plants11131739/s1, Figure S1 to S18. Figure S1. 1H NMR (CDCl3, 600 MHz) of compound (1). Figure S2. 13C NMR (CDCl3, 150 MHz) of compound (1). Figure S3. 3C (DEPT) NMR (CDCl3, 150 MHz) of compound (1). Figure S4. 1H-1H COSY NMR (CDCl3, 600 MHz) of compound (1). Figure S5. 1H-13C (HSQC) NMR (CDCl3, 600 MHz) of compound (1). Figure S6. 1H-13C (HMBC) NMR (CDCl3, 600 MHz) of compound (1). Figure S7. 1H NMR (CDCl3, 600 MHz) of salviandulin (2). Figure S8. 13C NMR (CDCl3, 150 MHz) of salviandulin (2). Figure S9. 3C (DEPT) NMR (CDCl3, 150 MHz) of salviandulin (2). Figure S10. 1H-1H COSY NMR (CDCl3, 600 MHz) of salviandulin (2). Figure S11. 1H-13C (HSQC) NMR (CDCl3, 600 MHz) of salviandulin (2). Figure S12. 1H-13C (HMBC) NMR (CDCl3, 600 MHz) of salviandulin (2). Figure S13. 1H NMR (CD3COCD3, 600 MHz) of eupatorin (3). Figure S14. 13C NMR (CD3COCD3, 150 MHz) of eupatorin (3). Figure S15. 3C (DEPT) NMR (CD3COCD3, 150 MHz) of eupatorin (3). Figure S16. 1H-1H COSY NMR (CD3COCD3, 600 MHz) of eupatorin (3) Figure S17. 1H-13C (HSQC) NMR (CD3COCD3, 600 MHz) of eupatorin (3). Figure S18. 1H-13C (HMBC) NMR (CD3COCD3, 600 MHz) of eupatorin (3).

Author Contributions

M.D.P.-G., M.G.-C., D.O.S.-S. and M.H.-R. conceived and designed the experiments; M.D.P.-G., M.G.-C., E.J.-F., P.Á.-F., A.Z. and R.C.-S. analyzed the chemical and biological data; M.D.P.-G., D.C.R.-R. and E.A.B.-G. performed the experiments; M.D.P.-G., M.G.-C. and D.O.S.-S. wrote/edited the paper. All authors have read and agreed to the published version of the manuscript.

Funding

This research received no external funding.

Acknowledgments

The authors acknowledge the Mexican Social Security Institute F-2021-1702-022. This research did not receive any specific grant from funding agencies from the public, commercial, or not-for-profit sectors. This work is a bachelor’s thesis developed by Dionisio Celerino Román Ramos.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Ceravantes, G.E.; García, G.R.; Salazar, S.P.M. Importance of Staphylococcus aureus methicillin resistant in-hospital and acquired in the community. Mex. J. Clin. Pathol. Lab. Med. 2014, 61, 196–204. [Google Scholar]
  2. CONAMED-PAHO Bulletin; 2018; Volume 3, p. 17.
  3. World Health Organization (WHO). WHO Publishes List of Bacteria for Which New Antibiotics Are Urgently Needed. 2017. Available online: http://www.who.int/news-room/detail/27-02-2017-who-publishes-list-of-bacteria-for-which-new-antibiotics-are-urgently-needed (accessed on 1 June 2017).
  4. Harris, L.G.; Foster, S.J.; Richards, R.G. An introduction to Staphylococcus aureus, and techniques for identifying and quantifying S. aureus adhesins in relation to adhesion to biomaterials: Review. Eur. Cells Mater. 2002, 4, 100–120. [Google Scholar] [CrossRef] [PubMed]
  5. Sordé, R.; Pahissa, A.; Rello, J. Management of refractory Pseudomonas aeruginosa infection in cystic fibrosis. Infect. Drug. Resis. 2011, 4, 31–41. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  6. Mendoza, A.H.; García, O.S.; Mata, R.R.; Pichardo, M.C.M.; Aguirre, V.M.B. The enzyme glycogen synthase kinase 3 regulates the activity of transcription factors NF-kB and creb during the inflammatory response caused by Staphylococcus aureus. J. Biochem. Educ. 2018, 36, 111–117. [Google Scholar]
  7. Elinav, E.; Nowarski, R.; Thaiss, C.A.; Hu, B.; Jin, C.; Flavell, R.A. Inflammation-induced cancer: Crosstalk between tumours, immune cells and microorganisms. Nat. Rev. Cancer 2013, 13, 759–771. [Google Scholar] [CrossRef]
  8. Suarez, C.; Godiol, F. Antibioticos betalactamicos. Enferm. Infecc. Y Microbiol. Clin. 2009, 27, 116–129. [Google Scholar] [CrossRef]
  9. Loza, E. NSAIDs in clinical practice: What you need to know. Inf. Ter. Nat. Sys. Health 2011, 35, 88–95. [Google Scholar]
  10. Guzmán, S.H.; Maldonado, R.S.; Díaz, H.M.; González, C.M. Medicinal Plants the Reality of an Ancestral Tradition; National Institute of Agricultural and Livestock Forestry Research Regional Research Center Bajío Celaya Experimental Field Center: Guanajuato, Mexico, 2017.
  11. López, L.T. Anti-inflammatory medicinal plants used in the treatment of rheumatism. Phytotherapy 2003, 22, 118–122. [Google Scholar]
  12. Jash, S.K.; Gorai, D.; Roy, R. Salvia genus and triterpenoids. Int. J. Pharm. Sci. Res. 2016, 7, 4710. [Google Scholar]
  13. Walker, J.B.; Elisens, W.J. A revision of Salvia section Heterosphace (Lamiaceae) in western North America. AIDS Contrib. Bot. 2001, 19, 571–589. [Google Scholar]
  14. Ramamoorthy, T.P.; Elliott, M. Lamiaceae of Mexico: Diversity, distribution, endemism and evolution. In Biological Diversity of Mexico: Origins and Distribution; Ramamoorthy, T.P., Bye, R., Lot, A., Fa, J., Eds.; Institute of Biology, National Autonomous University of Mexico: Coyoacán, Mexico, 1993; pp. 501–526. [Google Scholar]
  15. Villaseñor, J.L. The vascular plant genera of the flora of Mexico. Bol. Soc. Bot. Méx. 2004, 75, 105–135. [Google Scholar]
  16. Cornejo, T.G.; Ibarra, M.G. Diversity and distribution of the genus Salvia (Lamiaceae) in Michoacán, Mexico. Rev. Mex. De Biodivers. 2011, 82, 1279–1296. [Google Scholar]
  17. González, G.J.G.; Bedolla, G.B.Y.; Cornejo, T.G.; Fernández, A.J.L.; Fragoso, M.I.; García, P.M.D.R.; Harley, R.M.; Klitgaard, B.; Martínez, G.M.J.; Wood, J.R.I.; et al. Richness and distribution of Salvia subg. Calosphace (Lamiaceae). Int. J. Plant Sci. 2020, 181, 831–856. [Google Scholar] [CrossRef]
  18. Baricevic, D.; Sosa, S.; Della, L.R.; Tubaro, A.; Simonovska, B.; Krasna, A.; Zupancic, A. Topical anti-inflammatory activity of Salvia officinalis L. leaves: The relevance of ursolic acid. J. Ethnopharmacol. 2001, 75, 125–132. [Google Scholar] [CrossRef]
  19. Todorov, S.; Philianos, S.; Petkov, V.; Harvala, C.; Zamfirova, R.; Olimpiou, H. Experimental pharmacological study of three species from genus Salvia. Acta Physiol. Pharmacol. Bul. 1984, 10, 13–20. [Google Scholar]
  20. Ulubelen, A. Cardioactive and antibacterial terpenoids from some Salvia species. Phytochemistry 2003, 64, 395–399. [Google Scholar] [CrossRef]
  21. Gaitán, F.I.C. Activity of Twelve Native Guatemalan Plants against Sporothrix Schenckii. Biology Chemestry Thesis. University of San Carlos de Guatemala: Guatemala, 2005. Available online: http://biblioteca.usac.edu.gt/tesis/06/06_2289.pdf (accessed on 18 April 2022).
  22. Monroy, O.C.; Castillo, E.P. Medicinal Plants Used in the State of Morelos, 2nd ed.; Autonomous University of the State of Morelos: Cuernavaca, Mexico, 2007; p. 405. [Google Scholar]
  23. Del Cid, A.N.E. Activity of Seventeen Extracts of Twelve Native Guatemalan Plants against Fonsecaea Pedrosoi; USAC: Guatemala, 2005. [Google Scholar]
  24. Vázquez, G.D.; Castro, R.A.E. Medicinal uses of the Labiatae family in Chiapas, Mexico. Ethnobiology 2002, 2, 19–31. [Google Scholar]
  25. Arellano, R.B. Medicinal Ethnobotany of the Me’phaa Culture in La Ciénega, Municipality of Malinaltepec, Guerrero, Mexico; UAGro: Chilpancingo, Mexico, 2017. [Google Scholar]
  26. Palacios, V.M.C. Inhibition of the Growth of Gardnerella Vaginalis by Six Medicinal Plants of the Southwestern Guatemalan Flora; University of San Carlos de Guatemala: Guatemala, 2004; Available online: https://biblioteca-farmacia.usac.edu.gt/Tesis/QB782.pdf (accessed on 20 April 2022).
  27. Ortega, A.; Cárdenas, J.; Toscano, A.; Maldonado, E.; Aumelas, A.; Van, C.M.R.; Jankowski, C. Salviandulines A and B. Two secoclerodane diterpenoids from Salvia lavanduloides. Phytochemistry 1991, 30, 3357–3360. [Google Scholar] [CrossRef]
  28. Maldonado, E.; Cardenas, J.; Salazar, B.; Ortega, A.; Jankowski, C.K.; Van Calsteren, M.R. Salvianduline C, a 5, 6-secoclerodane diterpenoid from Salvia lavanduloides. Phytochemistry 1992, 31, 217–220. [Google Scholar] [CrossRef]
  29. González, C.M.M.; Ramos, V.C.S.; Serrano, V.R.; Pérez, G.C.; Sánchez, M.E.; Pérez, G.S. Anti-inflammatory activity of standardized dichloromethane extract of Salvia connivens on macrophages stimulated by LPS. Pharm. Biol. 2017, 55, 1467–1472. [Google Scholar] [CrossRef] [Green Version]
  30. Bisio, A.; Damonte, G.; Fraternale, D.; Giacomelli, E.; Salis, A.; Romussi, G.; Cafaggi, S.; Ricci, D.; De Tommasi, N. Phytotoxic clerodane diterpenes from Salvia miniata Fernald (Lamiaceae). Phytochemistry 2011, 72, 265–275. [Google Scholar] [CrossRef] [PubMed]
  31. Nagao, T.; Abe, F.; Kinjo, J.; Okabe, H. Antiproliferative constituents in plants 10. Flavones from the leaves of Lantana montevidensis B RIQ. and consideration of structure–activity relationship. Biol. Pharm. Bull. 2002, 25, 875–879. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  32. Argueta, A.; Cano, A.L.M.; Rodarte, G.M.E. Atlas of the Plants of Traditional Mexican Medicine; Instituto Nacional Indigenista: Mexico City, Mexico, 1994.
  33. Wu, Y.B.; Ni, Z.Y.; Shi, Q.W.; Dong, M.; Kiyota, H.; Gu, Y.C.; Cong, B. Constituents from Salvia species and their biological activities. Chem. Rev. 2012, 112, 5967–6026. [Google Scholar] [CrossRef] [PubMed]
  34. Lucas, S. The pharmacology of indomethacin. Headache J. Head Face Pain 2016, 56, 436–446. [Google Scholar] [CrossRef]
  35. Laavola, M.; Nieminen, R.; Yam, M.F.; Sadikun, A.; Asmawi, M.Z.; Basir, R.; Welling, J.; Vapaatalo, H.; Korhonen, R.; Moilanen, E. Flavonoids eupatorin and sinensetin present in Orthosiphon stamineus leaves inhibit inflammatory gene expression and STAT1 activation. Planta Med. 2012, 78, 779–786. [Google Scholar] [CrossRef]
  36. De La Cruz, S.N.G.; Gómez, R.A.; Alvarez, F.P.; Ventura, Z.E.; Pérez, G.M.D.; Avilés, F.M.; Gutiérrez, R.A.S.; González, C.M. Antibacterial activity of Morinda citrifolia Linneo sedes against Methicillin Resistant Staphylococcus spp. Microb. Pathog. 2019, 128, 347–353. [Google Scholar] [CrossRef]
  37. Monterrosas, B.N.; Ocampo, M.L.A.; Jiménez, F.E.; Jiménez, A.A.R.; Zamilpa, A.; Gonzalez, C.M.; Tortoriello, J.; Herrera, R.M. Anti-inflammatory activity of different Agave plants and the compound Cantalasaponin-1. Molecules 2013, 18, 8136–8146. [Google Scholar] [CrossRef]
  38. Salinas, S.D.O.; Herrera, R.M.; Pérez, S.; Jiménez, F.E.; Zamilpa, A. Anti-inflammatory activity of hautriwaic acid isolated from Dodonaea viscosa leaves. Molecules 2012, 17, 4292–4299. [Google Scholar] [CrossRef]
Plants 11 01739 g001 550
Figure 1. Chemical structures of compounds isolated from the SlD-3 fraction.
Figure 1. Chemical structures of compounds isolated from the SlD-3 fraction.
Plants 11 01739 g001
Plants 11 01739 g002 550
Figure 2. Protocol used to obtain all extracts, fractions, and compounds of Salvia lavanduloides. 15,16-epoxy-10-β-hydroxy-neo-cleroda-3,7,13(16),14-tetraene-17,12R:18,19-diolide (1), salviandulin A and (2) eupatorin (3).
Figure 2. Protocol used to obtain all extracts, fractions, and compounds of Salvia lavanduloides. 15,16-epoxy-10-β-hydroxy-neo-cleroda-3,7,13(16),14-tetraene-17,12R:18,19-diolide (1), salviandulin A and (2) eupatorin (3).
Plants 11 01739 g002
Table
Table 1. NMR spectroscopic data for compounds 1 and 2 (600 MHz, CDCl3, δ ppm).
Table 1. NMR spectroscopic data for compounds 1 and 2 (600 MHz, CDCl3, δ ppm).
Positionδ1H (J in Hz)
1		δ13C
1		δ1H (J in Hz)
2		δ13C
2
1a
b		1.79, dd(8.0, 14.3)
1.72, ddd(4.0, 11.3, 15.4)		25.36.13, ddd(1.1, 2.5, 10.2)129.7
2a
b		2.40, dd(4.0, 8.0)
2.41, dd(4.0, 8.0)		22.96.98, dddd(0.7, 2.5, 5.5, 10.2)149.3
3a
b		6.79, dd(3.3, 3.6)135.63.01, dd(br, 5.1, 20.1)
2.76, ddd(2.5, 6.9, 20.5)		23.2
4 131.843.07, d(br, 6.9)43.3
5 44.6 51.6
6a
b		2.79, dd(4.4, 20.5)
2.33, d(1.8, 20.5)		39.102.69, dd(8.0, 14.6)
2.33, d(4.4, 14.6)		39.1
76.42, dd(3.3, 4.0)133.15.63, d(4.4, 8.0)67.3
8 134.8 124.9
9 40.82 154.8
10 71.7 197.4
11a
b		2.66, dd (2.2, 15.4)
1.70, dd(12.4, 15.4)		38.52.74, m
2.51, dd(3.6, 17.9)		37.5
125.03, dd(1.83, 12.4)71.915.3, d(3.6, 11)71.0
13 123.7 123.5
146.63 d(br, 1.4)109.036.42, d(br, 1.1)108.5
157.69, dd(1.4, 1.8)144.07.42, dd(1.8, 1.8)143.9
167.79, d(br, 0.7)140.587.47, dd(0.7, 0.7)140.1
17 167.8 163.0
18 168.96 176.5
19a
b		4.29, d(8.0)
4.33, d(8.0)		73.354.74, d(8.8)
3.84, d(8.8)		73.6
201.10, s29.202.13, s20.4
MeCO2- 2.03, s170.2, 21.1
Table
Table 2. NMR spectroscopic data for eupatorin (3, 600 MHz, CD3COCD3, δ ppm, J in Hz).
Table 2. NMR spectroscopic data for eupatorin (3, 600 MHz, CD3COCD3, δ ppm, J in Hz).
Positionδ1H (in ppm, J in Hz)
3		δ 13C
3
2 165.3
36.67(s)104.2
4 183.5
5 154.0
6 133.4
7 160.0
86.86(s)92.0
9 153.5
10 106.3
1′ 124.4
2′7.48 (d, 1.4)112.6
3′ 148.0
4′ 152.1
5′7.55 (dd, 1.4, 8.4)113.7
6′7.10 (d, 8.4)119.6
6-OCH33.79 (s)56.3
7-OCH33.92(s)56.8
4′-OCH33.97(s)60.5
Table
Table 3. Antimicrobial activity of Salvia lavanduloides extracts.
Table 3. Antimicrobial activity of Salvia lavanduloides extracts.
Extracts
μg/mL
BacteriumSl-HexSl-AcOEtSl-DControl (+)Control (−)
Sa>200200100---†
SaR>200200100---†
Se-1100<25100---†
Se-2>200200200---†
Se-3>200200200---†
Sh>200200200---†
Ef<25<25<25---†
Pa<25<25<25---†
Sd<25<25<25---†
Ca<25<25<25††
(---): No growth; (†) growth. Staphylococcus aureus ATCC 29213 (Sa), methicillin-resistant Staphylococcus aureus ATCC 42310 (SaR), Staphylococcus epidermis ATCC 35984 (Se-1), Staphylococcus epidermis ATCC 12228 (Se-2), Staphylococcus epidermis ATCC 1042 (Se-3), Staphylococcus haemolitycus ATCC 1165 (Sh), Enterococcus faecalis ATCC 29212 (Ef), Gram-negative bacteria Pseudomonas aeruginosa ATCC 27853 (Pa), Salmonella dublin NTCC 9676 (Sd), and the fungus Candida albicans ATCC 10231 (Ca). n-hexane (Sl-Hex), ethyl acetate (Sl-AcOEt), and dichloromethane (Sl-D).
Table
Table 4. Antimicrobial activity of Salvia lavanduloides fractions and compounds.
Table 4. Antimicrobial activity of Salvia lavanduloides fractions and compounds.
Fractions μg/mLμg/mL Compounds
BacteriumSlD-2SlD-3123Control (+)Control (−)
Sa100200161616---†
SaR50200161616---†
Se-150<25161616---†
Se-2100200161616---†
Se-3100200161616---†
Sh100100161616---†
Ef<25<25<2<2<2---†
Pa<25<25<2<2<2---†
Sd<25>200<216<2---†
Ca>200>200<21616††
(---): does not grow; (†) growth. Staphylococcus aureus ATCC 29213 (Sa), methicillin-resistant Staphylococcus aureus ATCC 42310 (SaR), Staphylococcus epidermis ATCC 35984 (Se-1), Staphylococcus epidermis ATCC 12228 (Se-2), Staphylococcus epidermis ATCC 1042 (Se-3), Staphylococcus haemolitycus ATCC 1165 (Sh), Enterococcus faecalis ATCC 29212 (Ef), Gram-negative bacteria Pseudomonas aeruginosa ATCC 27853 (Pa), Salmonella dublin NTCC 9676 (Sd), and the fungus Candida albicans ATCC 10231 (Ca). SlD-2 and SlD-3 fractions. 15,16-epoxy-10-β-hydroxy-neo-cleroda-3,7,13(16),14-tetraene-17,12R:18,19-diolide (1), salviandulin A (2), and eupatorin (3).
Table
Table 5. Anti-inflammatory activity of Salvia lavanduloides.
Table 5. Anti-inflammatory activity of Salvia lavanduloides.
Treatments
(1.0 mg/ear)		Edema (mg)Inflammation
Inhibition (%)
VEH11.76 ± 1.400
INDO2.72 ± 0.7576.87
Extracts
Sl-Hex4.46 ± 2.0462.02
Sl-D6.50 ± 1.2344.73
Sl-AcOEt6.45 ± 1.1045.15
Fractions
SlD-29.73 ± 2.1417.23
SlD-34.56 ± 1.4061.22
Compounds
Salviandulin A (2)3.50 ± 0.6770.24
Eupatorin (3)3.25 ± 1.01 *72.45
Extracts, fractions, and compounds S. lavanduloides, TPA (acetone); INDO, indomethacin. Data show mean ± S.E.M. of seven animals. * p < 0.05 indicates statistically significant differences using ANOVA followed by the Dunnett test compared with VEH-Tween group.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Share and Cite

MDPI and ACS Style

González-Cortazar, M.; Salinas-Sánchez, D.O.; Herrera-Ruiz, M.; Román-Ramos, D.C.; Zamilpa, A.; Jiménez-Ferrer, E.; Ble-González, E.A.; Álvarez-Fitz, P.; Castrejón-Salgado, R.; Pérez-García, M.D. Eupatorin and Salviandulin-A, with Antimicrobial and Anti-Inflammatory Effects from Salvia lavanduloides Kunth Leaves. Plants 2022, 11, 1739. https://doi.org/10.3390/plants11131739

AMA Style

González-Cortazar M, Salinas-Sánchez DO, Herrera-Ruiz M, Román-Ramos DC, Zamilpa A, Jiménez-Ferrer E, Ble-González EA, Álvarez-Fitz P, Castrejón-Salgado R, Pérez-García MD. Eupatorin and Salviandulin-A, with Antimicrobial and Anti-Inflammatory Effects from Salvia lavanduloides Kunth Leaves. Plants. 2022; 11(13):1739. https://doi.org/10.3390/plants11131739

Chicago/Turabian Style

González-Cortazar, Manasés, David Osvaldo Salinas-Sánchez, Maribel Herrera-Ruiz, Dionisio Celerino Román-Ramos, Alejandro Zamilpa, Enrique Jiménez-Ferrer, Ever A. Ble-González, Patricia Álvarez-Fitz, Ricardo Castrejón-Salgado, and Ma. Dolores Pérez-García. 2022. "Eupatorin and Salviandulin-A, with Antimicrobial and Anti-Inflammatory Effects from Salvia lavanduloides Kunth Leaves" Plants 11, no. 13: 1739. https://doi.org/10.3390/plants11131739

APA Style

González-Cortazar, M., Salinas-Sánchez, D. O., Herrera-Ruiz, M., Román-Ramos, D. C., Zamilpa, A., Jiménez-Ferrer, E., Ble-González, E. A., Álvarez-Fitz, P., Castrejón-Salgado, R., & Pérez-García, M. D. (2022). Eupatorin and Salviandulin-A, with Antimicrobial and Anti-Inflammatory Effects from Salvia lavanduloides Kunth Leaves. Plants, 11(13), 1739. https://doi.org/10.3390/plants11131739

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop