GC-MS identification of bactericidal and antibiofilm compounds from the essential oils of Lippia multiflora Moldenke (Verbeaceae) flowers: a promising study to combat bacterial resistance

GC-MS identification of bactericidal and antibiofilm compounds from the essential oils of Lippia multiflora Moldenke (Verbeaceae) flowers: a promising study to combat bacterial resistance | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article GC-MS identification of bactericidal and antibiofilm compounds from the essential oils of Lippia multiflora Moldenke (Verbeaceae) flowers: a promising study to combat bacterial resistance Ablassé Rouamba, Moussa Compaoré, Yahaya Zoungrana, Eli Compaoré, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7414241/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 28 Nov, 2025 Read the published version in Scientific Reports → Version 1 posted 18 You are reading this latest preprint version Abstract Background: Bacterial resistance to conventional antibiotic therapy has become a global health crisis. Bacterial biofilm formation allows bacteria to develop resistance to antibiotics and the host immune system. This study aims to contribute for fighting against bacterial resistance through the inhibition of bacterial biofilm formation by Lippia multiflora flowers essential oils. Methodology: The sensitivity of Staphylococcus aureus and Streptococcus mutans to the essential oils was measured on the Luria Bertani-Agar mediumby using the antibiotic susceptibility test. The anti-biofilm potential of the essential oil was evaluated by t he crystal violet method. The antioxidant potential of the essential oils was assessed by the DPPH method. The phytochemical profile was determined by gas chromatography-mass spectrometry (GC/MS) analysis. Results: The essential oils showed higher growth inhibition diameters of S. aureus (28.38±0.23 mm) and S. mutans (24.5±0.25 mm) than reference antibiotics such as gentamicin 30 mg (13.5 ± 0.07 mm on S. aureus ) and Oleandomycin 10μg (16.5 ± 0.05 mm on S. mutans ).The essential oil was also more active in inhibiting biofilm formation than salicylic acid with inhibition percentages ranged from 60 ± 0.00% on S. aureus and 71.60 ± 0.70% on S. mutans . Moreover, the essential oils exhibited good antioxidant activity with anti-DPPH inhibitory concentration ranged from 0.23 ± 0.02 µg/ùL. GC-MS analysis of the essential oils led to the identification of 22 terpens compounds with 7 major compounds such as β-caryophyllene (43.55%), germacrene D (16.44%), elemol (10.33%), eucalyptol (6.41%), humulene (3.28%), α-phellandrene (2.68%) and β-ocimene (2.62%). Conclusion: These results showed that the essential oils of L. multiflora flowers are a promising candidate in the fight against bacterial resistance. Health sciences/Health care Biological sciences/Microbiology bacterial resistance biofilm essential oil Lippia multiflora gas chromatography-mass spectrometry Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Introduction Antibiotic resistance is a major public health problem [ 1 ]. It occurs when bacteria evolve to become less sensitive or even insensitive to drugs designed to kill them or inhibit their growth. This phenomenon makes bacterial infections more difficult to treat, increasing the risk of complications, serious illness, and death [ 2 ]. Several resistance mechanisms are developed by bacteria, including the enzymatic deactivation of antibiotics, the rejection of antibiotics by reflux pumps and the reduction of antibiotic permeability due to the biofilm formation [ 3 ]. Biofilm is a form of resistance, particularly in the context of microbial infections [ 4 ]. Biofilms, which are communities of microorganisms (bacteria, fungi, etc.) encased in a protective matrix, exhibit increased resistance to antibiotics, disinfectants and the immune system compared to planktonic microorganisms [ 5 ]. Biofilm tolerance has been attributed to many variables, including slow growth, the presence of an exopolysaccharide matrix that could block the diffusion of antibiotics and the presence of undiscovered resistance mechanisms. Bacteria in biofilms can be up to 1,000 times more resistant to antibiotics than those in a planktonic state [ 6 ]. This resistance results from the close proximity of the organisms, which promotes communication and the transfer of resistance genes as well as an increase in the virulence factors synthesis. Streptococcus mutans , a bacterium found in the mouth, plays a key role in the formation of dental plaque and tooth decay [ 7 ]. It forms biofilms, a complex bacterial community that provides increased protection against host immune defenses and antimicrobial treatments, thereby increasing its resistance. S. mutan s produces glucosyltransferases that convert sugar into glucans, promoting the bacteria's adhesion to tooth surfaces and plaque formation [ 8 ]. The ability of S. mutans to form biofilms is a major factor in its virulence and resistance, making it an important target for the development of strategies to prevent and treat oral diseases. S. aureus is a Gram-positive bacterium that can cause a variety of infections, ranging from mild skin infections to more serious infections such as pneumonia or endocarditis [ 9 ]. Its ability to form biofilms and its resistance to antibiotics are two key factors that contribute to its pathogenicity and difficulty in treatment. Biofilm-associated infections can be chronic, recurrent and lead to significant morbidity and mortality. Current research focuses on understanding the mechanisms of biofilm formation, the antibiotic resistance, and developing new therapeutic strategies to combat bacterial infections, particularly by targeting biofilms [ 10 ]. Promising approaches include the development of new antibiotics such as the use of phages or the inhibition of biofilm formation by natural compounds. In the biofilm state, bacteria use general mechanisms of resistance to antimicrobial agents, including integrons, chemical modification of antibiotics, damage caused by antibiotics, reduction of their penetration, efflux pump, target protection, as well as other resistance mechanisms [ 4 ]. There are several strategies to eradicate adhesion and prevent bacterial accumulation in the structure of biofilms. In fact, the objective of these methods is to restore cells from the biofilm state to the planktonic state, decreasing bacterial virulence and increasing sensitivity to antibiotics. The most promising therapeutic methods are the development of antibiofilm phytomolecules [ 11 ]. Lippia multiflora is a aromatic plant belong the verbenacea family which the leaves and flowers were consumed as tea in the West Africa regions [ 12 ]. Its aerial part essential oils have proven antibacterial properties, reported by several scientific studies [ 13 ]. It is traditionally used in West Africa for various ailments, and its active compounds, such as linalool, 1,8-cineole, and citral, contribute to its antimicrobial activity [ 14 ]. Previous studies showed that L. multiflora essential oils possesses antimicrobial activities on different bacteria, including Gram-postive and Gram-negative strains. It has been successfully tested against bacteria such as S. aureus and Escherichia coli , two common pathogens. These antibacterial properties are attributed to the presence of certain chemical compounds such as monoterpenes and phenols in the essential oils [ 14 ]. However, data on the antibiofilm potentiality of the flower’s essential oils against multiresistant bacteria like S. mutans and S. aureus are not available. This study aims to contribute for fighting against bacterial resistance by inhibiting S. mutans and S. aureus biofilm formation by using the essential oils of L. multiflora flowers. Materials and Methods Chemical 2,2-diphenyl-1-picrylhydrazyl (DPPH), Dimethyl Sulfoxide, Luria Bertani (LB), P-iodonitrotetrazolium (INT), Agar, Ascorbic acid and Salicylic acid were purchased by Sigma Aldrich, Germany. Methanol, Acetic acid were purchased from Pro Labo (France) Plant material collection The flowers of wild L. multiflora were harvested in September 2022 at Loumbila, a locality located 25 km from Ouagadougou (12°31’5.39˝N / 1°22’8.39ʺW). Plant collect is complied with ethical and legal guidelines at the institutional, national, and international levels, including obtaining permits, respecting private property, avoiding harm to ecosystems, and ensuring the sustainability of wild populations according to the International Treaty on Plant Genetic Resources for Food and Agriculture guidelines. A specimen of the leafy stem was collected and brought to the plant biology and ecology laboratory for identification by Professor Amadé Ouédraogo, full professor, botanist of the University Joseph KI-ZERBO, and a herbarium was deposited at the UFR/SVT herbarium under the identification code IC-922. The flowers were dried under laboratory conditions (away from light, room temperature) and pulverized using a grinder. The powder obtained was stored in a refrigerator at 4°C until the extraction of essential oils. Essential oils extraction The essential oils were extracted by hydrodistillation using a Clevenger-type apparatus as described previously with slight modifications [15] 100g of the flower powder were placed in a glass flask and 1500 mL of distilled water were added. The mixture was brought to a boil using a hot plate. The vapors containing the essential oils were condensed in a refrigerated column and the distillate (oil + water) was collected in a separating funnel. The distillate was filtered in the presence of sodium sulfate to remove residual traces of water. The essential oils obtained were stored at 4°C away from light for the phytochemical and pharmacological investigations. Organoleptic characteristics and physicochemical parameters determination The color and the appearance of the essential oil were determined by direct observation in the visible, and the sensory was done by inhalation. The relative density was determined by using a pycnometer at 20°C. Results were compared to AFNOR normalization. DPPH quenching assay The antioxidant property of the essential oils was measured by using the DPPH method [16]. A series of seven (07) successive dilutions was carried out from the sample solutions of 1% essential oils diluted in methanol. For each concentration, four repeated experiments were carried out by mixing 100 µL of sample and 200 µL of DPPH (20 mg/L in methanol). After 15 minutes of incubation, the optical densities were recorded at 517 nm using an Epoch UV-Visible spectrophotometer. Ascorbic acid was used as a reference compound. The 50% inhibitory concentrations of the DPPH radical were calculated using the inhibition percentages. Antibiotic Susceptibility Test The properties of the essential oils to inhibit bacterial growth were evaluated by measuring diffusion diameters on agar medium [17]. Bacterial inocula were plated in petri dishes containing LB agar at 1.5 × 10 8 CFU/mL. Whatman paper discs impregnated with 10 µL of 100% essential oils were placed on the agar. Two antibiotics (cefotaxime 30 μg and oleandomycin 10 μg) were used as positive controls. After incubation at 37°C for 24 h, the inhibition zones were photographed and the diameters of the inhibition zones were measured. Determination of minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) In order to determine the non-bactericidal and the non-bacteriostatic concentration of the essential oils for the antibiofilm investigation, the MIC and the MBC of the essential oils were determined as described previously [18]. The MIC was determined by using the serial liquid dilution process. A half-fold dilution series of the essential oils in LB containing 1% DMSO was performed in wells of a sterile 96-well plate, and 30 µL of bacterial inoculum (1.5 × 10 8 CFU) were added to each well. The plate was incubated for 18 h at 37°C, and 30 µL of 0.2 g/mL p-iodonitrotetrazolium (INT) were added and incubated for an additional 30 minutes. The MIC was deduced from the first well in the series devoid of bacterial growth, marked by the absence of the pink INT color. For the determination of the CMB, samples were taken from the wells without bacterial growth at the INT and placed in petri dishes containing LB-agar growth medium. The petri dishes were incubated for 24 h at 37 °C and the CMB was determined from the dish containing the lowest concentration of essential oils without visible bacteria. Antibiofilm assay The ability of the essential oils to inhibit bacterial biofilm was measured by using the crystal violet method [19]. Each bacterial inoculum (1.5 × 10 8 CFU) was incubated in the presence of non-bacteriostatic and non-bactericidal concentrations of the essential oils (0.05%) or salicylic acid (100 µg/mL) for 18 h at 37 °C. Planktonic bacteria were removed and the formed biofilms were fixed with 100 µL of methanol for 15 min. The methanol was then removed and 100 µL of crystal violet were added. After 30 min of incubation at 37 °C, the excess crystal violet was washed off with distilled water and the crystal violet fixed by the biofilm was dissolved with 100 µL of acetic acid. The optical density were measured at 590 nm by using a UV-Visible spectrometer and the anti-biofilm activity was expressed as percentage inhibition relative to a control without extract. GC - MS analysis of essential oils The GC-MS analysis of the essential oils was performed with an Agilent 8860 GC-FID instrument equipped with a DB Wax column (PEG, 60m x 250 µm x 0.25µm) associated with an MSD (5977 GC-MSD Agilent) mass spectrometer as performed in the literature with slight modifications [20] . The transfer-line temperature and the ionization voltage were 250 °C and 70 eV respectively. The oven conditions are summarized in the table 1. Table 1: Oven conditions Rate (°C/min) Temperature (°C) Retention time (min) Flow time (min) Initial - 40 2 2 Ramp 10 260 10 34 The carrier gas was air (400.0 ml/min), H 2 (30ml/min), injection volume 1 µL, injection mode Split 1/3 (MSD); 2/3 (FID), concentration of sample 20µL/mL. The compounds were identified based on a comparison of their retention time and their mass spectra with those of standard compounds by using the National Institut of Standard and Technology databases. The CAS numbers of the identified compounds were verified. Statistical analysis Data were expressed as the mean value of several independent repetitive experiments (n ≥ 4) ± standard deviation. The degree of significance between the results was verified by One Way ANOVA analysis of variance followed by Newman Keuls post test. A statistical difference was observed at P > 0.05 . Results Organoleptic characteristics and the physicochemical properties of the essential oils The organoleptic characteristics and physicochemical properties of the essential oils were shown in Table 2 . The essential oils of L. multiflora flowers presented a mobile liquid appearance, which is in accordance with AFNOR data. The slight differences observed in color and odor compared to AFNOR data would be due to the extraction conditions as well as abiotic and biotic factors. Table 2 Organoleptic characteristics and the physicochemical parameters determination Parameters Essential oil AFNOR NFT 75.11 Appearance Mobile liquid Mobile liquid, clear Color Pale yellow Amber yellow-greenish yellow Smell Dew characteristics Minty dew Relative density (g/mL) 0.9432 - Anti-DPPH activity The antioxidant potential of the essential oils was evaluated by measuring their ability to trap the DPPH radical and data were showed in the Fig. 1 . The essential oils showed a higher anti-DPPH activity than ascorbic acid (P < 0.001) with a 50% inhibitory concentration ranged from 0.23 ± 0.02 µg/mL. This highest antioxidant potential of the essential oils may due to a synergistic effect between the different volatile reducing phytomolecules. Antibiotic Susceptibility activity The sensitivity of the resistant strains S. mutans and S. aureus to the essential oils was studied on LB-agar solid medium. The bacterial inhibition zones were photographed (Fig. 2 ) and the diameters of the inhibition zones were showed in Table 3 . The essential oils exhibited higher growth inhibitory activities of S. mutans ATCC 25175 and S. aureus ATCC43300 than those of the reference antibiotics with inhibition diameters ranged from 24.50 ± 0.25 mm and 28.38 ± 0.23 mm, respectively. S. aureus was more sensitive to the essential oils than S. mutans. Table 3 Inhibition diameters of essential oils Essential oils / antibiotics Inhibition diameters (mm) S. aureus ATCC 43300 S. mutans ATCC 25175 Essential oils 100% 28.38 ± 0.23 a 24.50 ± 0.25 a Cefotaxime 30 µg 13.50 ± 0.07 b Inactive Oleandomycine 10µg Inactive 16.50 ± 0.05 b Values in each column with different superscript letters were statically different (P < 0.05). Determination of minimal inhibitory and minimal bactericidal concentrations The data of the minimum inhibitory and bactericidal concentrations of the essential oils were presented in Table 4 . The essential oils of L. multiflora flowers showed a minimum inhibitory concentration of 0.10 ± 0.00% and a minimum bactericidal concentration greater than 0.10 ± 0.00% on both the two bacteria. This subsequently allows choosing a non-bacteriostatic and non-bactericidal concentration of 0.05% for the measurement of the antibiofilm potential. Table 4 minimal inhibitory and minimal bactericidal concentrations S. aureus ATCC 43300 S. mutans ATCC 25175. MIC MBC MIC MBC Essential oils (%) 0.10 ± 0.00 > 0.10 ± 0.01 0.10 ± 0.00 > 0.10 ± 0.00 Salycilic acid (µg/mL) > 100.00 ± 0.00 > 100.00 ± 0.01 > 100.00 ± 0.01 > 100.00 ± 0.01 MIC: minimal inhibitory concentration; MBC: minimal bactericidal concentration Antibiofilm activity The antibiofilm potential of the essential oils was presented in the Fig. 3 . The essential oils showed higher inhibitory activity on the biofilm formation in S. aureu s and S. mitans than the reference salicylic acid (P < 0.001) with inhibition percentages of around 60% and 70% respectively. The essential oils inhibited the biofilm formation of S. mutans much more than those of S. aureus . These results suggested a very uninteresting antibiofilm potential of the essential oils L. multiflora flowers. Chemical analysis of the essential oils The phytomolecules of the essential oils were identified and quantified by GC/MS by using the chromatogram of the essential oils (Fig. 4 ) and the chromatogram of standards. The identified compounds along with their retention time and their proportion are presented in the Table 5 . The chemical structures of these compounds are shown in the Fig. 5 . A total of 22 terpenes and derivatives compounds were identified with seven (7) major compounds caryophyllene (43.55%), germacrene D (16.44%), elemol (10.33%), eucalyptol (6.41%), humulene (3.28%), α-phellandrene (2.68%) and β-ocimene (2.62%). These compounds are divided into 4 groups according to the number of carbons and the degree of oxidation such as hydrocarbon monoterpenes, oxygenated monoterpenes, hydrocarbon sesquiterpenes and oxygenated sesquiterpenes. Table 5 Chemical profiling of the Essential oils RT; Retention Time; CAS: Chemical Abstracts Service; k: Retention Factor; * Majority compounds N° Compounds name CAS number Prob (%) k RT (min) % Hydrocarbon monoterpens 1 α-pinene 80-56-8 24.20 1025 6.48 0.79 2 sabinene 3387-41-5 50.60 1124 8.03 1.54 3 α-phellandrene 99-83-2 45.20 1169 8.73 2.68* 4 D-limonene 138-86-3 33.30 1203 9.25 1.08 5 β-ocimene 3779-61-1 16.60 1252 9.98 2.62* 6 p-cymene 99-87-6 21.60 1275 10.32 0.81 Oxygen monoterpens 7 eucalyptol 470-82-6 89.40 1218 9.47 6.41* 8 camphor 76-22-2 40.90 1539 13.92 1.20 Hydrocarbon sesquiterpens 9 β-copaene 18252-44-3 16.40 1595 14.64 0.52 10 α-copaene 3856-25-5 36.60 1509 13.55 1.22 11 β-elemene 515-13-9 17.60 1606 14.77 1.03 12 β-caryophyllene 87-44-5 40,50 1626 15,00 43.55* 13 alloaromadendrene 25246-27-9 12.50 1671 15.54 1,08 14 humulene 6753-98-6 45.10 1694 15.82 3.28* 15 germacrene D 23986-74-5 48.40 1736 16.29 16.44* 16 bicyclogermacrene 24703-35-3 34.00 1758 16.54 0.57 17 δ-cadinene 483-76-1 27.20 1774 16.73 0.98 Oxygen sesquiterpens 18 caryophyllene oxide 1139-30-6 66.80 2020 19.34 0.80 19 γ-eudesmol 1209-71-8 54.40 2187 20.96 0.96 20 α-eudesmol 473-16-5 53.90 2244 21.49 1.01 21 β-eudesmol 473-15-4 76.40 2255 21.59 1.09 22 elemol 639-99-6 36.80 2091 20.05 10.33* Discussion The essential oils possess antibacterial properties due to their chemical composition, particularly their volatile compounds, which can inhibit bacterial growth and multiplication [ 13 , 14 ]. These oils can be used to prevent infections, help cure certain conditions and strengthen the immune system. Indeed, essential oils have the properties of inhibiting bacterial growth by preventing the multiplication of bacteria, the formation of spores, the synthesis of their toxins and the organization of their plasma membrane [ 21 ]. They also have the properties of enhancing the action of antibiotics and neutralizing the mechanism of bacterial resistance [ 22 ]. Strategies targeting the formation of S. mutans and S. aureus biofilms, particularly by interfering with the metabolism of exopolysaccharides (EPS), offer promising pathways for preventing infections caused by these bacteria. These strategies aim to disrupt EPS synthesis, degrade existing EPS, or interfere with the regulatory systems controlling their production. S. mutans uses glucosyltransferases (Gtf) to synthesize glucans from sucrose [ 8 ]. Inhibiting these enzymes, either directly or by targeting their regulatory pathways could reduce the amount of produced EPS, thereby impeding the biofilm formation. EPS, particularly glucans, form a matrix that binds S. mutans cells to each other and to the tooth surface, contributing to the biofilm formation and protection. Enzymes such as glucanases can degrade this matrix, disrupting the biofilm and potentially making the bacterium more susceptible to other treatments [ 23 ]. The essential oils of L. multiflora flowers showed in this study an interesting inhibitory properties against the biofilm formation in S. aureus and S. mutans . These essential oils would inhibit EPS synthesis by trapping Gtfs involved in EPS synthesis or by targeting Gtf regulatory pathways. β-caryophyllene was identified in this current study as the major compound of the L. multiflora flower essential oils that would be responsible for the antibiofilm properties of essential oils demonstrated. Indeed, molecular docking studies identified the β-caryophyllene as a potent inhibitor of enterococcal surface protein (Esp) involved in the initial adhesion and the formation of biofilm [ 24 ]. Futhermore, the essential oils significantly inhibited the growth of both bacteria on LB-agar medium with more pronounced inhibitory activities than the reference antibiotics cefotaxime 30 µg and oleandomycin 10 µg. β-caryophyllene would thought to be responsible for this bactericidal activity of the essential oils. Indeed, in previous studies, β-caryophyllene showed higher inhibitory activity on the growth of S aureus than the antibiotic kanamycin [ 25 ]. In addition, β-caryophyllene showed antibacterial activity against S. mutans , especially at concentrations above 0.078% [ 26 ]. It has been also suggested that β-caryophyllene could be an alternative to chlorhexidine, a commonly used antiseptic, in the prevention and treatment of periodontal diseases [ 27 ]. Previous research has shown that antioxidant molecules can modulate biofilm formation by interfering with signaling and communication mechanisms between bacteria, thus preventing the biofilm formation. For example, antioxidants like ascorbic acid degraded the extracellular matrix of the biofilm, making bacteria more vulnerable to antimicrobial treatments [ 28 ]. By weakening the biofilm, antioxidants can potentially increase the effectiveness of antibiotics, allowing better penetration and more effective action against bacteria. In this current study, the essential oils showed strong antioxidant potential against the DPPH radical, higher than ascorbic acid. The antioxidant potential of the essential oils may be partly responsible for its antibiofilm properties demontrated, and could be benefit in the treatment of numerous infectious diseases associated to an oxidative stress. Conclusion The essential oils of L. multiflora flowers showed very interesting bactericidal and antibiofilm potentials on two multiresistant bacterial strains S. aureus ATCC43300 and S. mutans ATCC 25175 as well as a powerful antioxidant property. These combined pharmacological potentials make these essential oils a potential candidate for the development of a phytomedicament against infectious diseases caused by multiresistant germs. Abbreviations CFU : Colony Forming Units MBC: Minimal Bactericidal Concentration MIC: Minimal Inhibitory Concentration DPPH: 2,2-diphenyl-1-picrylhydrazyl EPS : Exopolysaccharides GC-MS: Gaz Chromatography-Mass Sperctrometry Gtf: G lucosyltransferases IC 50 : Inhibitory concentration 50% INT: iodonitrotetrazolium LB: Luria Bertani SD: Standard deviation UFR/SVT : Unité de Formation et de Recherche en Sciences de la Vie et de la Terre Declarations Ethics approval and consent to participate Not applicable Consent for publication Not applicable Availability of data and materials All data were presented in this current manuscript Conflicts of Interest All authors declare no conflicts of interest Funding Not applicable Author’s contribution A.R wrote original draft, methodology and data curation. M.C performed the GC/MS analysis. Y.Z; E.C performed the antibacterial investigation and data curation. M.K reviewed the manuscript and check the grammar. 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Cite Share Download PDF Status: Published Journal Publication published 28 Nov, 2025 Read the published version in Scientific Reports → Version 1 posted Editorial decision: Revision requested 21 Oct, 2025 Reviews received at journal 19 Oct, 2025 Reviews received at journal 12 Oct, 2025 Reviews received at journal 10 Oct, 2025 Reviews received at journal 10 Oct, 2025 Reviewers agreed at journal 06 Oct, 2025 Reviews received at journal 06 Oct, 2025 Reviews received at journal 04 Oct, 2025 Reviewers agreed at journal 02 Oct, 2025 Reviewers agreed at journal 01 Oct, 2025 Reviewers agreed at journal 01 Oct, 2025 Reviewers agreed at journal 30 Sep, 2025 Reviewers agreed at journal 30 Sep, 2025 Reviewers invited by journal 30 Sep, 2025 Editor assigned by journal 29 Sep, 2025 Editor invited by journal 26 Aug, 2025 Submission checks completed at journal 23 Aug, 2025 First submitted to journal 23 Aug, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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14:25:31","extension":"html","order_by":100,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":113560,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-7414241/v1/1c1751a465fa78e72c6c012d.html"},{"id":93409139,"identity":"31a5e041-a771-4cf8-b190-55bf2e8a1621","added_by":"auto","created_at":"2025-10-13 14:17:28","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":71166,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eAnti-DPPH activity of the essential oils\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-7414241/v1/9fea985239a3e5de3f1356e3.png"},{"id":93410987,"identity":"05ced549-87ba-4b0d-84ff-1948f389f301","added_by":"auto","created_at":"2025-10-13 14:33:28","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":353847,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003ePhoto of inhibition diameters\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eA:\u003c/strong\u003eEssential oils 100%;\u003cstrong\u003e B:\u003c/strong\u003eOleandomycine 10μg;\u003cstrong\u003e C: \u003c/strong\u003eCefotaxime 30µg; \u003cstrong\u003eE:\u003c/strong\u003eEssential oils 100%;\u003cstrong\u003e F: \u003c/strong\u003eCefotaxime 30µg\u003cstrong\u003e; G:\u003c/strong\u003eOleandomycine 10μg\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-7414241/v1/669b1d79105453225eb4d24c.png"},{"id":93410499,"identity":"d86ae0d8-84b7-4fdc-a8c2-59ab3e483c54","added_by":"auto","created_at":"2025-10-13 14:25:28","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":143828,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eAntibiofilm activity of the essential oils; \u003c/strong\u003eValues are expressed as Mean ± SD (n=6).\u003cstrong\u003e \u003c/strong\u003e\u003csup\u003e\u003cstrong\u003e***\u003c/strong\u003e\u003c/sup\u003e\u003cem\u003eP \u0026lt;0.001 significant difference from \u003c/em\u003e\u0026nbsp;the salicylic acid\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-7414241/v1/9e188528ca11562414b473c6.png"},{"id":93409144,"identity":"e0f313fa-b43a-4eac-8d1a-dacde0ebf713","added_by":"auto","created_at":"2025-10-13 14:17:29","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":109428,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eThe essential oils chromatogram\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-7414241/v1/09597fbbcb9f17cf6cc22b17.png"},{"id":93409148,"identity":"52bbf776-3db8-4946-b1f3-1c8a6e43eb2c","added_by":"auto","created_at":"2025-10-13 14:17:29","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":59653,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eThe chemical structures of the identified compounds\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-7414241/v1/288faebdc7ff10b39aa565cb.png"},{"id":97178219,"identity":"7d77a0f8-3f76-4f66-aff0-89d7a8224f94","added_by":"auto","created_at":"2025-12-01 16:00:54","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2014912,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7414241/v1/bcc4793e-c07c-4d35-8088-94cc78da17c5.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"GC-MS identification of bactericidal and antibiofilm compounds from the essential oils of Lippia multiflora Moldenke (Verbeaceae) flowers: a promising study to combat bacterial resistance","fulltext":[{"header":"Introduction","content":"\u003cp\u003eAntibiotic resistance is a major public health problem [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. It occurs when bacteria evolve to become less sensitive or even insensitive to drugs designed to kill them or inhibit their growth. This phenomenon makes bacterial infections more difficult to treat, increasing the risk of complications, serious illness, and death [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. Several resistance mechanisms are developed by bacteria, including the enzymatic deactivation of antibiotics, the rejection of antibiotics by reflux pumps and the reduction of antibiotic permeability due to the biofilm formation [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Biofilm is a form of resistance, particularly in the context of microbial infections [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. Biofilms, which are communities of microorganisms (bacteria, fungi, etc.) encased in a protective matrix, exhibit increased resistance to antibiotics, disinfectants and the immune system compared to planktonic microorganisms [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Biofilm tolerance has been attributed to many variables, including slow growth, the presence of an exopolysaccharide matrix that could block the diffusion of antibiotics and the presence of undiscovered resistance mechanisms. Bacteria in biofilms can be up to 1,000 times more resistant to antibiotics than those in a planktonic state [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. This resistance results from the close proximity of the organisms, which promotes communication and the transfer of resistance genes as well as an increase in the virulence factors synthesis.\u003c/p\u003e\u003cp\u003e\u003cem\u003eStreptococcus mutans\u003c/em\u003e, a bacterium found in the mouth, plays a key role in the formation of dental plaque and tooth decay [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. It forms biofilms, a complex bacterial community that provides increased protection against host immune defenses and antimicrobial treatments, thereby increasing its resistance. \u003cem\u003eS. mutan\u003c/em\u003es produces glucosyltransferases that convert sugar into glucans, promoting the bacteria's adhesion to tooth surfaces and plaque formation [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. The ability of \u003cem\u003eS. mutans\u003c/em\u003e to form biofilms is a major factor in its virulence and resistance, making it an important target for the development of strategies to prevent and treat oral diseases. \u003cem\u003eS. aureus\u003c/em\u003e is a Gram-positive bacterium that can cause a variety of infections, ranging from mild skin infections to more serious infections such as pneumonia or endocarditis [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Its ability to form biofilms and its resistance to antibiotics are two key factors that contribute to its pathogenicity and difficulty in treatment. Biofilm-associated infections can be chronic, recurrent and lead to significant morbidity and mortality.\u003c/p\u003e\u003cp\u003eCurrent research focuses on understanding the mechanisms of biofilm formation, the antibiotic resistance, and developing new therapeutic strategies to combat bacterial infections, particularly by targeting biofilms [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Promising approaches include the development of new antibiotics such as the use of phages or the inhibition of biofilm formation by natural compounds. In the biofilm state, bacteria use general mechanisms of resistance to antimicrobial agents, including integrons, chemical modification of antibiotics, damage caused by antibiotics, reduction of their penetration, efflux pump, target protection, as well as other resistance mechanisms [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. There are several strategies to eradicate adhesion and prevent bacterial accumulation in the structure of biofilms. In fact, the objective of these methods is to restore cells from the biofilm state to the planktonic state, decreasing bacterial virulence and increasing sensitivity to antibiotics. The most promising therapeutic methods are the development of antibiofilm phytomolecules [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e].\u003c/p\u003e\u003cp\u003e\u003cem\u003eLippia multiflora\u003c/em\u003e is a aromatic plant belong the verbenacea family which the leaves and flowers were consumed as tea in the West Africa regions [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Its aerial part essential oils have proven antibacterial properties, reported by several scientific studies [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. It is traditionally used in West Africa for various ailments, and its active compounds, such as linalool, 1,8-cineole, and citral, contribute to its antimicrobial activity [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. Previous studies showed that \u003cem\u003eL. multiflora\u003c/em\u003e essential oils possesses antimicrobial activities on different bacteria, including Gram-postive and Gram-negative strains. It has been successfully tested against bacteria such as \u003cem\u003eS. aureus\u003c/em\u003e and \u003cem\u003eEscherichia coli\u003c/em\u003e, two common pathogens. These antibacterial properties are attributed to the presence of certain chemical compounds such as monoterpenes and phenols in the essential oils [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. However, data on the antibiofilm potentiality of the flower\u0026rsquo;s essential oils against multiresistant bacteria like \u003cem\u003eS. mutans\u003c/em\u003e and \u003cem\u003eS. aureus\u003c/em\u003e are not available. This study aims to contribute for fighting against bacterial resistance by inhibiting \u003cem\u003eS. mutans\u003c/em\u003e and \u003cem\u003eS. aureus\u003c/em\u003e biofilm formation by using the essential oils of \u003cem\u003eL. multiflora\u003c/em\u003e flowers.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cp\u003e\u003cstrong\u003eChemical\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e2,2-diphenyl-1-picrylhydrazyl (DPPH), Dimethyl Sulfoxide, Luria Bertani (LB), P-iodonitrotetrazolium (INT), Agar, Ascorbic acid and Salicylic acid were purchased by Sigma Aldrich, Germany. \u0026nbsp;Methanol, Acetic acid were purchased from Pro Labo (France)\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePlant material collection\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe flowers of\u003cem\u003e\u0026nbsp;\u003c/em\u003ewild\u0026nbsp;\u003cem\u003eL. multiflora\u003c/em\u003e were harvested in September 2022\u0026nbsp;at\u0026nbsp;Loumbila, a locality located 25 km from Ouagadougou (12\u0026deg;31\u0026rsquo;5.39˝N / 1\u0026deg;22\u0026rsquo;8.39ʺW).\u0026nbsp;Plant collect is complied with ethical and legal guidelines at the institutional, national, and international levels, including obtaining permits, respecting private property, avoiding harm to ecosystems, and ensuring the sustainability of wild populations according to the International Treaty on Plant Genetic Resources for Food and Agriculture guidelines.\u0026nbsp;A specimen of the leafy stem was collected and brought to the plant biology and ecology laboratory for identification by Professor Amad\u0026eacute;\u0026nbsp;Ou\u0026eacute;draogo, full professor, botanist of the University Joseph KI-ZERBO,\u0026nbsp;and a herbarium was deposited at the UFR/SVT herbarium under the identification\u0026nbsp;code\u0026nbsp;IC-922. The flowers were dried under laboratory conditions (away\u0026nbsp;from light, room temperature) and pulverized using a grinder. The powder obtained was stored in a refrigerator at 4\u0026deg;C until the extraction of essential oils.\u0026nbsp;\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEssential oils extraction\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe essential oils were extracted by hydrodistillation using a Clevenger-type apparatus as described previously with slight modifications [15] 100g of the flower powder were placed in a glass flask and 1500 mL of distilled water were added. The mixture was brought to a boil using a hot plate. The vapors containing the essential oils were condensed in a refrigerated column and the distillate (oil + water) was collected in a separating funnel. The distillate was filtered in the presence of sodium sulfate to remove residual traces of water. The essential oils obtained were stored at 4\u0026deg;C away from light for the phytochemical and pharmacological investigations.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eOrganoleptic characteristics and \u0026nbsp;physicochemical parameters determination\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;The color and the appearance of the essential oil were determined by direct observation in the visible, and the sensory was done by inhalation. The relative density was determined by using a pycnometer at 20\u0026deg;C. Results were compared to AFNOR normalization.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDPPH quenching assay\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe antioxidant property of the essential oils was measured by using the DPPH method [16]. A series of seven (07) successive dilutions was carried out from the sample solutions of 1% essential oils diluted in methanol. For each concentration, four repeated experiments were carried out by mixing 100 \u0026micro;L of sample and 200 \u0026micro;L of DPPH (20 mg/L in methanol). After 15 minutes of incubation, the optical densities were recorded at 517 nm using an Epoch UV-Visible spectrophotometer. Ascorbic acid was used as a reference compound. The 50% inhibitory concentrations of the DPPH radical were calculated using the inhibition percentages.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAntibiotic Susceptibility Test\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe properties of the essential oils to inhibit bacterial growth were evaluated by measuring diffusion diameters on agar medium [17]. Bacterial inocula were plated in petri dishes containing LB agar at 1.5 \u0026times; 10\u003csup\u003e8\u0026nbsp;\u003c/sup\u003eCFU/mL. Whatman paper discs impregnated with 10 \u0026micro;L of 100% essential oils were placed on the agar. Two antibiotics (cefotaxime 30 \u0026mu;g and oleandomycin 10 \u0026mu;g) were used as positive controls. After incubation at 37\u0026deg;C for 24 h, the inhibition zones were photographed and the diameters of the inhibition zones were measured.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDetermination of minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn order to determine the non-bactericidal and the non-bacteriostatic concentration of the essential oils for the antibiofilm investigation, the MIC and the MBC of the essential oils were determined as described previously [18].\u0026nbsp;The MIC\u0026nbsp;was determined\u0026nbsp;by\u0026nbsp;using the\u0026nbsp;serial\u0026nbsp;liquid dilution\u0026nbsp;process. A half-fold dilution series of\u0026nbsp;the essential oils\u0026nbsp;in LB containing 1% DMSO was performed in wells of a sterile 96-well plate, and 30 \u0026micro;L of bacterial inoculum (1.5 \u0026times; 10\u003csup\u003e8\u003c/sup\u003e CFU) were\u0026nbsp;added to each well. The plate was incubated for 18 h at 37\u0026deg;C,\u0026nbsp;and 30 \u0026micro;L of 0.2 g/mL p-iodonitrotetrazolium (INT) were\u0026nbsp;added and incubated for an additional 30 minutes. The MIC was deduced from the first well in the series devoid of bacterial growth, marked by the absence of the pink INT color.\u003c/p\u003e\n\u003cp\u003eFor the determination of the CMB, samples were taken from the wells without bacterial growth at the INT and placed in petri dishes containing LB-agar growth medium. The petri dishes were incubated for 24 h at 37 \u0026deg;C and the CMB was determined from the dish containing the lowest concentration of essential oils without visible bacteria.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAntibiofilm assay\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe ability of the essential oils to inhibit bacterial biofilm was measured by using the crystal violet method [19]. Each bacterial inoculum (1.5 \u0026times; 10\u003csup\u003e8\u003c/sup\u003e CFU) was incubated in the presence of \u0026nbsp; non-bacteriostatic and \u0026nbsp; non-bactericidal concentrations of the essential oils (0.05%) or salicylic acid (100 \u0026micro;g/mL) for 18 h at 37 \u0026deg;C. Planktonic bacteria were removed and the formed biofilms were fixed with 100 \u0026micro;L of methanol for 15 min. The methanol was then removed and 100 \u0026micro;L of crystal violet were added. After 30 min of incubation at 37 \u0026deg;C, the excess crystal violet was washed off with distilled water and the crystal violet fixed by the biofilm was dissolved with 100 \u0026micro;L of acetic acid. The optical density were measured at 590 nm by using a UV-Visible spectrometer and the anti-biofilm activity was expressed as percentage inhibition relative to a control without extract.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eGC\u003c/strong\u003e\u003cstrong\u003e-\u003c/strong\u003e\u003cstrong\u003eMS analysis of essential oils\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe GC-MS analysis of the essential oils was performed with an Agilent 8860 GC-FID instrument equipped with a DB Wax column (PEG, 60m x 250 \u0026micro;m x 0.25\u0026micro;m) associated with an MSD (5977 GC-MSD Agilent) mass spectrometer as performed in the literature with slight modifications [20] . The transfer-line temperature and the ionization voltage were 250 \u0026deg;C and 70 eV respectively. The oven conditions are summarized in the table 1.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 1: Oven conditions\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"624\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 76px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 104px;\"\u003e\n \u003cp\u003eRate (\u0026deg;C/min)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 132px;\"\u003e\n \u003cp\u003eTemperature (\u0026deg;C)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 154px;\"\u003e\n \u003cp\u003eRetention time (min)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 158px;\"\u003e\n \u003cp\u003eFlow time (min)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 76px;\"\u003e\n \u003cp\u003eInitial\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 104px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 132px;\"\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 154px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 158px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 76px;\"\u003e\n \u003cp\u003eRamp\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 104px;\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 132px;\"\u003e\n \u003cp\u003e260\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 154px;\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 158px;\"\u003e\n \u003cp\u003e34\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eThe carrier gas was air (400.0 ml/min), H\u003csub\u003e2\u003c/sub\u003e (30ml/min), injection volume 1 \u0026micro;L, injection mode \u0026nbsp; \u0026nbsp;Split 1/3 (MSD); 2/3 (FID), concentration of sample 20\u0026micro;L/mL. The compounds were identified based on a comparison of their retention time and their mass spectra with those of standard compounds\u0026nbsp;by\u0026nbsp;using the\u0026nbsp;National Institut of Standard and Technology\u0026nbsp;databases. The CAS numbers of the identified compounds were verified.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eStatistical analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eData were expressed as the mean value of several independent repetitive experiments (n\u0026nbsp;\u0026ge;\u0026nbsp;4) \u0026plusmn; standard deviation. The degree of significance between the results was verified by One Way ANOVA\u0026nbsp;analysis of variance followed by Newman Keuls post test. A statistical difference was observed at \u003cem\u003eP \u0026gt; 0.05\u003c/em\u003e.\u003c/p\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec14\" class=\"Section2\"\u003e\n \u003ch2\u003eOrganoleptic characteristics and the physicochemical properties of the essential oils\u003c/h2\u003e\n \u003cp\u003eThe organoleptic characteristics and physicochemical properties of the essential oils were shown in Table \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e. The essential oils of \u003cem\u003eL. multiflora\u003c/em\u003e flowers presented a mobile liquid appearance, which is in accordance with AFNOR data. The slight differences observed in color and odor compared to AFNOR data would be due to the extraction conditions as well as abiotic and biotic factors.\u003c/p\u003e\n \u003cp\u003e\u003c/p\u003e\n \u003ctable id=\"Tab2\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eOrganoleptic characteristics and the physicochemical parameters determination\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eParameters\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eEssential oil\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eAFNOR NFT 75.11\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAppearance\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMobile liquid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMobile liquid, clear\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eColor\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ePale yellow\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAmber yellow-greenish yellow\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSmell\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eDew characteristics\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMinty dew\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eRelative density (g/mL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.9432\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n \u003cp\u003e\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec15\" class=\"Section2\"\u003e\n \u003ch2\u003eAnti-DPPH activity\u003c/h2\u003e\n \u003cp\u003eThe antioxidant potential of the essential oils was evaluated by measuring their ability to trap the DPPH radical and data were showed in the Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e. The essential oils showed a higher anti-DPPH activity than ascorbic acid (P\u0026thinsp;\u0026lt;\u0026thinsp;0.001) with a 50% inhibitory concentration ranged from 0.23\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02 \u0026micro;g/mL. This highest antioxidant potential of the essential oils may due to a synergistic effect between the different volatile reducing phytomolecules.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec16\" class=\"Section2\"\u003e\n \u003ch2\u003eAntibiotic Susceptibility activity\u003c/h2\u003e\n \u003cp\u003eThe sensitivity of the resistant strains \u003cem\u003eS. mutans\u003c/em\u003e and \u003cem\u003eS. aureus\u003c/em\u003e to the essential oils was studied on LB-agar solid medium. The bacterial inhibition zones were photographed (Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e) and the diameters of the inhibition zones were showed in Table \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e. The essential oils exhibited higher growth inhibitory activities of \u003cem\u003eS. mutans\u003c/em\u003e ATCC 25175 and \u003cem\u003eS. aureus\u003c/em\u003e ATCC43300 than those of the reference antibiotics with inhibition diameters ranged from 24.50\u0026thinsp;\u0026plusmn;\u0026thinsp;0.25 mm and 28.38\u0026thinsp;\u0026plusmn;\u0026thinsp;0.23 mm, respectively. \u003cem\u003eS. aureus\u003c/em\u003e was more sensitive to the essential oils than \u003cem\u003eS. mutans.\u003c/em\u003e\u003c/p\u003e\n \u003cp\u003e\u003c/p\u003e\n \u003ctable id=\"Tab3\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eInhibition diameters of essential oils\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\" colspan=\"2\"\u003e\n \u003cp\u003eEssential oils / antibiotics\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\" colspan=\"4\"\u003e\n \u003cp\u003eInhibition diameters (mm)\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003cth align=\"left\" colspan=\"2\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eS. aureus\u003c/em\u003e ATCC 43300\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eS. mutans\u003c/em\u003e ATCC 25175\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEssential oils 100%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\" colspan=\"2\"\u003e\n \u003cp\u003e28.38\u0026thinsp;\u0026plusmn;\u0026thinsp;0.23\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\" colspan=\"2\"\u003e\n \u003cp\u003e24.50\u0026thinsp;\u0026plusmn;\u0026thinsp;0.25\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\" colspan=\"1\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCefotaxime 30 \u0026micro;g\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\" colspan=\"2\"\u003e\n \u003cp\u003e13.50\u0026thinsp;\u0026plusmn;\u0026thinsp;0.07\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\" colspan=\"2\"\u003e\n \u003cp\u003eInactive\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\" colspan=\"1\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eOleandomycine 10\u0026micro;g\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\" colspan=\"2\"\u003e\n \u003cp\u003eInactive\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\" colspan=\"2\"\u003e\n \u003cp\u003e16.50\u0026thinsp;\u0026plusmn;\u0026thinsp;0.05\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\" colspan=\"1\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n \u003cp\u003e\u003c/p\u003e\n \u003cp\u003eValues in each column with different superscript letters were statically different (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05).\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec17\" class=\"Section2\"\u003e\n \u003ch2\u003eDetermination of minimal inhibitory and minimal bactericidal concentrations\u003c/h2\u003e\n \u003cp\u003eThe data of the minimum inhibitory and bactericidal concentrations of the essential oils were presented in Table \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e. The essential oils of \u003cem\u003eL. multiflora\u003c/em\u003e flowers showed a minimum inhibitory concentration of 0.10\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00% and a minimum bactericidal concentration greater than 0.10\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00% on both the two bacteria. This subsequently allows choosing a non-bacteriostatic and non-bactericidal concentration of 0.05% for the measurement of the antibiofilm potential.\u003c/p\u003e\n \u003cp\u003e\u003c/p\u003e\n \u003ctable id=\"Tab4\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eminimal inhibitory and minimal bactericidal concentrations\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eS. aureus\u003c/em\u003e ATCC 43300\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eS. mutans\u003c/em\u003e ATCC 25175.\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMIC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMBC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMIC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMBC\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEssential oils (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.10\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026gt;\u0026thinsp;0.10\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.10\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026gt;\u0026thinsp;0.10\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSalycilic acid (\u0026micro;g/mL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026gt;\u0026thinsp;100.00\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026gt;\u0026thinsp;100.00\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026gt;\u0026thinsp;100.00\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026gt;\u0026thinsp;100.00\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003ctfoot\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"5\"\u003eMIC: minimal inhibitory concentration; MBC: minimal bactericidal concentration\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tfoot\u003e\n \u003c/table\u003e\n \u003cp\u003e\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec18\" class=\"Section2\"\u003e\n \u003ch2\u003eAntibiofilm activity\u003c/h2\u003e\n \u003cp\u003eThe antibiofilm potential of the essential oils was presented in the Fig. \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e. The essential oils showed higher inhibitory activity on the biofilm formation in \u003cem\u003eS. aureu\u003c/em\u003es and \u003cem\u003eS. mitans\u003c/em\u003e than the reference salicylic acid (P\u0026thinsp;\u0026lt;\u0026thinsp;0.001) with inhibition percentages of around 60% and 70% respectively. The essential oils inhibited the biofilm formation of \u003cem\u003eS. mutans\u003c/em\u003e much more than those of \u003cem\u003eS. aureus\u003c/em\u003e. These results suggested a very uninteresting antibiofilm potential of the essential oils \u003cem\u003eL. multiflora\u003c/em\u003e flowers.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec19\" class=\"Section2\"\u003e\n \u003ch2\u003eChemical analysis of the essential oils\u003c/h2\u003e\n \u003cp\u003eThe phytomolecules of the essential oils were identified and quantified by GC/MS by using the chromatogram of the essential oils (Fig. \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e) and the chromatogram of standards. The identified compounds along with their retention time and their proportion are presented in the Table \u003cspan class=\"InternalRef\"\u003e5\u003c/span\u003e. The chemical structures of these compounds are shown in the Fig. \u003cspan class=\"InternalRef\"\u003e5\u003c/span\u003e. A total of 22 terpenes and derivatives compounds were identified with seven (7) major compounds caryophyllene (43.55%), germacrene D (16.44%), elemol (10.33%), eucalyptol (6.41%), humulene (3.28%), \u0026alpha;-phellandrene (2.68%) and \u0026beta;-ocimene (2.62%). These compounds are divided into 4 groups according to the number of carbons and the degree of oxidation such as hydrocarbon monoterpenes, oxygenated monoterpenes, hydrocarbon sesquiterpenes and oxygenated sesquiterpenes.\u003c/p\u003e\n \u003cp\u003e\u003c/p\u003e\n \u003ctable id=\"Tab5\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 5\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003e\u003cstrong\u003eChemical profiling of the Essential oils\u003c/strong\u003e RT; Retention Time; CAS: Chemical Abstracts Service; k: Retention Factor; \u003csup\u003e*\u003c/sup\u003eMajority compounds\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eN\u0026deg;\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eCompounds name\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eCAS number\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eProb (%)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003ek\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eRT (min)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e%\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\" colspan=\"6\"\u003e\n \u003cp\u003eHydrocarbon monoterpens\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026alpha;-pinene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e80-56-8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e24.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1025\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e6.48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.79\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003esabinene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e3387-41-5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e50.60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1124\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e8.03\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.54\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026alpha;-phellandrene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e99-83-2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e45.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1169\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e8.73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2.68*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eD-limonene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e138-86-3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e33.30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1203\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e9.25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.08\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026beta;-ocimene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e3779-61-1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e16.60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1252\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e9.98\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2.62*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ep-cymene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e99-87-6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e21.60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1275\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e10.32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.81\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\" colspan=\"7\"\u003e\n \u003cp\u003e\u003cstrong\u003eOxygen monoterpens\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eeucalyptol\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e470-82-6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e89.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1218\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e9.47\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e6.41*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ecamphor\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e76-22-2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e40.90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1539\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e13.92\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.20\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\" colspan=\"7\"\u003e\n \u003cp\u003e\u003cstrong\u003eHydrocarbon sesquiterpens\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026beta;-copaene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e18252-44-3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e16.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1595\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e14.64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.52\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026alpha;-copaene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e3856-25-5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e36.60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1509\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e13.55\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.22\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026beta;-elemene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e515-13-9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e17.60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1606\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e14.77\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.03\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e12\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026beta;-caryophyllene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e87-44-5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e40,50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1626\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e15,00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e43.55*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ealloaromadendrene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e25246-27-9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e12.50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1671\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e15.54\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1,08\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ehumulene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e6753-98-6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e45.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1694\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e15.82\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e3.28*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003egermacrene D\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e23986-74-5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e48.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1736\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e16.29\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e16.44*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ebicyclogermacrene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e24703-35-3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e34.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1758\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e16.54\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.57\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026delta;-cadinene\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e483-76-1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e27.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1774\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e16.73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.98\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\" colspan=\"7\"\u003e\n \u003cp\u003e\u003cstrong\u003eOxygen sesquiterpens\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ecaryophyllene oxide\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1139-30-6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e66.80\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2020\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e19.34\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.80\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e19\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026gamma;-eudesmol\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1209-71-8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e54.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2187\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e20.96\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e0.96\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026alpha;-eudesmol\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e473-16-5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e53.90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2244\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e21.49\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026beta;-eudesmol\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e473-15-4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e76.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2255\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e21.59\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e1.09\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e22\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eelemol\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e639-99-6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e36.80\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2091\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e20.05\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e10.33*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n \u003cp\u003e\u003c/p\u003e\n\u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe essential oils possess antibacterial properties due to their chemical composition, particularly their volatile compounds, which can inhibit bacterial growth and multiplication [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. These oils can be used to prevent infections, help cure certain conditions and strengthen the immune system. Indeed, essential oils have the properties of inhibiting bacterial growth by preventing the multiplication of bacteria, the formation of spores, the synthesis of their toxins and the organization of their plasma membrane [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. They also have the properties of enhancing the action of antibiotics and neutralizing the mechanism of bacterial resistance [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. Strategies targeting the formation of \u003cem\u003eS. mutans\u003c/em\u003e and \u003cem\u003eS. aureus\u003c/em\u003e biofilms, particularly by interfering with the metabolism of exopolysaccharides (EPS), offer promising pathways for preventing infections caused by these bacteria. These strategies aim to disrupt EPS synthesis, degrade existing EPS, or interfere with the regulatory systems controlling their production. \u003cem\u003eS. mutans\u003c/em\u003e uses glucosyltransferases (Gtf) to synthesize glucans from sucrose [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. Inhibiting these enzymes, either directly or by targeting their regulatory pathways could reduce the amount of produced EPS, thereby impeding the biofilm formation. EPS, particularly glucans, form a matrix that binds \u003cem\u003eS. mutans\u003c/em\u003e cells to each other and to the tooth surface, contributing to the biofilm formation and protection. Enzymes such as glucanases can degrade this matrix, disrupting the biofilm and potentially making the bacterium more susceptible to other treatments [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. The essential oils of \u003cem\u003eL. multiflora\u003c/em\u003e flowers showed in this study an interesting inhibitory properties against the biofilm formation in \u003cem\u003eS. aureus\u003c/em\u003e and \u003cem\u003eS. mutans\u003c/em\u003e. These essential oils would inhibit EPS synthesis by trapping Gtfs involved in EPS synthesis or by targeting Gtf regulatory pathways. β-caryophyllene was identified in this current study as the major compound of the \u003cem\u003eL. multiflora\u003c/em\u003e flower essential oils that would be responsible for the antibiofilm properties of essential oils demonstrated. Indeed, molecular docking studies identified the β-caryophyllene as a potent inhibitor of enterococcal surface protein (Esp) involved in the initial adhesion and the formation of biofilm [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eFuthermore, the essential oils significantly inhibited the growth of both bacteria on LB-agar medium with more pronounced inhibitory activities than the reference antibiotics cefotaxime 30 \u0026micro;g and oleandomycin 10 \u0026micro;g. β-caryophyllene would thought to be responsible for this bactericidal activity of the essential oils. Indeed, in previous studies, β-caryophyllene showed higher inhibitory activity on the growth of \u003cem\u003eS aureus\u003c/em\u003e than the antibiotic kanamycin [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. In addition, β-caryophyllene showed antibacterial activity against \u003cem\u003eS. mutans\u003c/em\u003e, especially at concentrations above 0.078% [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e]. It has been also suggested that β-caryophyllene could be an alternative to chlorhexidine, a commonly used antiseptic, in the prevention and treatment of periodontal diseases [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e].\u003c/p\u003e\u003cp\u003ePrevious research has shown that antioxidant molecules can modulate biofilm formation by interfering with signaling and communication mechanisms between bacteria, thus preventing the biofilm formation. For example, antioxidants like ascorbic acid degraded the extracellular matrix of the biofilm, making bacteria more vulnerable to antimicrobial treatments [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. By weakening the biofilm, antioxidants can potentially increase the effectiveness of antibiotics, allowing better penetration and more effective action against bacteria. In this current study, the essential oils showed strong antioxidant potential against the DPPH radical, higher than ascorbic acid. The antioxidant potential of the essential oils may be partly responsible for its antibiofilm properties demontrated, and could be benefit in the treatment of numerous infectious diseases associated to an oxidative stress.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThe essential oils of \u003cem\u003eL. multiflora\u003c/em\u003e flowers showed very interesting bactericidal and antibiofilm potentials on two multiresistant bacterial strains \u003cem\u003eS. aureus\u003c/em\u003e ATCC43300 and \u003cem\u003eS. mutans\u003c/em\u003e ATCC 25175 as well as a powerful antioxidant property. These combined pharmacological potentials make these essential oils a potential candidate for the development of a phytomedicament against infectious diseases caused by multiresistant germs.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003e\u003cstrong\u003eCFU\u003c/strong\u003e\u003cstrong\u003e:\u0026nbsp;\u003c/strong\u003eColony Forming Units\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMBC:\u0026nbsp;\u003c/strong\u003eMinimal Bactericidal Concentration\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMIC:\u0026nbsp;\u003c/strong\u003eMinimal Inhibitory Concentration\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDPPH:\u0026nbsp;\u003c/strong\u003e2,2-diphenyl-1-picrylhydrazyl\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEPS\u003c/strong\u003e\u003cstrong\u003e:\u0026nbsp;\u003c/strong\u003eExopolysaccharides\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eGC-MS:\u0026nbsp;\u003c/strong\u003eGaz Chromatography-Mass Sperctrometry\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eGtf: G\u003c/strong\u003elucosyltransferases\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eIC\u003csub\u003e50\u003c/sub\u003e:\u0026nbsp;\u003c/strong\u003eInhibitory concentration 50%\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eINT:\u0026nbsp;\u003c/strong\u003eiodonitrotetrazolium\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eLB:\u0026nbsp;\u003c/strong\u003eLuria Bertani\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSD:\u0026nbsp;\u003c/strong\u003eStandard deviation\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eUFR/SVT\u003c/strong\u003e: Unit\u0026eacute; de Formation et de Recherche en Sciences de la Vie et de la Terre\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll data were presented in this current manuscript\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflicts of Interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll authors declare no conflicts of interest\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor\u0026rsquo;s contribution\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eA.R\u0026nbsp;\u003c/strong\u003ewrote original draft, methodology and data curation. \u003cstrong\u003eM.C\u003c/strong\u003e performed the GC/MS analysis. \u003cstrong\u003eY.Z; E.C\u003c/strong\u003e performed the antibacterial investigation and data curation. M.K reviewed the manuscript and check the grammar. All authors read and validated the manuscript.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eBashir N, Dablool AS, Khan MI, Almalki MG, Ahmed A, Ahmad M, et al. Antibiotics resistance as a major public health concern_ A pharmaco-epidemiological study to evaluate prevalence and antibiotics susceptibility-resistance pattern of bacterial isolates from multiple teaching hospitals. J Infect Public Health [Internet]. 2023;16:61\u0026ndash;8. Available from: https://doi.org/10.1016/j.jiph.2023.09.019\u003c/li\u003e\n\u003cli\u003eSalam A, Al-amin Y, Salam MT, Pawar JS, Akhter N, Rabaan AA, et al. Antimicrobial Resistance : A Growing Serious Threat for Global Public Health. 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Antibiotics. 2024;13(625):1\u0026ndash;37. \u003c/li\u003e\n\u003cli\u003eAl-madboly LA, Aboulmagd A, El-salam MA, Kushkevych I, El-morsi RM. Microbial enzymes as powerful natural anti- biofilm candidates. Microb Cell Fact. 2024;23(343):1\u0026ndash;16. \u003c/li\u003e\n\u003cli\u003eKhyati K, Ishwerpreet KJ, Thomson S, Kashish M, Sunidhi B, Pranjali S, et al. Antibacterial and antibiofilm potential of Thuja orientalis L . extract targeting cariogenic Enterococcus faecalis ATCC 29212 : A combined in-vitro , in- silico study , and cytotoxicity screening. Arch Oral Biol. 2025;171:106107. \u003c/li\u003e\n\u003cli\u003eDahham SS, Tabana YM, Iqbal MA, Ahamed MBK, Ezzat MO, Majid ASA, et al. The Anticancer, Antioxidant and Antimicrobial Properties of the Sesquiterpene \u0026beta;-Caryophyllene from the Essential Oil of Aquilaria crassna. Molecules. 2015;20:11808\u0026ndash;29. \u003c/li\u003e\n\u003cli\u003eYoo H, Jwa S. Inhibitory effects of \u0026beta;-caryophyllene on Streptococcus mutans biofilm. Arch Oral Biol. 2018;88:42\u0026ndash;6. \u003c/li\u003e\n\u003cli\u003ePieri FA, Campos MSDC, Lobato LVR, Lobato MVR, Perciano PG, Vargas FS, et al. Use of \u0026beta; -caryophyllene to combat bacterial dental plaque formation in dogs. BMC Vet Res. 2016;12(216):1\u0026ndash;8. Available from: http://dx.doi.org/10.1186/s12917-016-0842-1\u003c/li\u003e\n\u003cli\u003ePandit S, Ravikumar V, Abdel-haleem AM, Derouiche A, Mokkapati VRSS, Sihlbom C, et al. Low Concentrations of Vitamin C Reduce the Synthesis of Extracellular Polymers and Destabilize Bacterial Biofilms. Front Microbiol. 2017;8(2599):1\u0026ndash;11. \u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"bacterial resistance, biofilm, essential oil, Lippia multiflora, gas chromatography-mass spectrometry","lastPublishedDoi":"10.21203/rs.3.rs-7414241/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7414241/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground:\u003c/strong\u003e Bacterial resistance to conventional antibiotic therapy has become a global health crisis. Bacterial biofilm formation allows bacteria to develop resistance to antibiotics and the host immune system. This study aims to contribute for fighting against bacterial resistance through the inhibition of bacterial biofilm formation by \u003cem\u003eLippia multiflora\u003c/em\u003e flowers essential oils.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethodology:\u003c/strong\u003e The sensitivity of \u003cem\u003eStaphylococcus aureus\u003c/em\u003e and \u003cem\u003eStreptococcus mutans\u003c/em\u003e to the essential oils was measured on the Luria Bertani-Agar mediumby using the antibiotic susceptibility test. The anti-biofilm potential of the essential oil was evaluated by t he crystal violet method. The antioxidant potential of the essential oils was assessed by the DPPH method. The phytochemical profile was determined by gas chromatography-mass spectrometry (GC/MS) analysis.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults:\u003c/strong\u003e The essential oils showed higher growth inhibition diameters of \u003cem\u003eS. aureus\u003c/em\u003e (28.38±0.23 mm) and \u003cem\u003eS. mutans\u003c/em\u003e (24.5±0.25 mm) than reference antibiotics such as gentamicin 30 mg (13.5 ± 0.07 mm on \u003cem\u003eS. aureus\u003c/em\u003e) and Oleandomycin 10μg (16.5 ± 0.05 mm on \u003cem\u003eS. mutans\u003c/em\u003e).The essential oil was also more active in inhibiting biofilm formation than salicylic acid with inhibition percentages ranged from 60 ± 0.00% on \u003cem\u003eS. aureus\u003c/em\u003e and 71.60 ± 0.70% on \u003cem\u003eS. mutans\u003c/em\u003e. Moreover, the essential oils exhibited good antioxidant activity with anti-DPPH inhibitory concentration ranged from 0.23 ± 0.02 µg/ùL. GC-MS analysis of the essential oils led to the identification of 22 terpens compounds with 7 major compounds such as β-caryophyllene (43.55%), germacrene D (16.44%), elemol (10.33%), eucalyptol (6.41%), humulene (3.28%), α-phellandrene (2.68%) and β-ocimene (2.62%).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusion:\u003c/strong\u003eThese results showed that the essential oils of \u003cem\u003eL. multiflora\u003c/em\u003e flowers are a promising candidate in the fight against bacterial resistance.\u003c/p\u003e","manuscriptTitle":"GC-MS identification of bactericidal and antibiofilm compounds from the essential oils of Lippia multiflora Moldenke (Verbeaceae) flowers: a promising study to combat bacterial resistance","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-10-13 14:17:23","doi":"10.21203/rs.3.rs-7414241/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-10-21T15:42:36+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-19T04:01:36+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-13T03:43:49+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-10T18:51:29+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-10T05:02:49+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"275455104405065467570888466441719905566","date":"2025-10-06T13:19:31+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-06T05:29:23+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-04T18:15:24+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"27195587918646926109162499128963595248","date":"2025-10-02T10:58:32+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"99031011549117219038521666005325974012","date":"2025-10-01T13:35:22+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"325176710579145269910896830745006984814","date":"2025-10-01T09:01:36+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"147038033087529100235958958124956719874","date":"2025-10-01T01:35:42+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"186004842334790590440090753532246024024","date":"2025-09-30T10:02:49+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-09-30T07:07:08+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-09-29T07:23:50+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2025-08-26T11:56:16+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-08-23T08:51:57+00:00","index":"","fulltext":""},{"type":"submitted","content":"Scientific Reports","date":"2025-08-23T08:48:42+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"ec3e5c5b-f56e-4892-b4b4-6cd6b2e67903","owner":[],"postedDate":"October 13th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[{"id":56111807,"name":"Health sciences/Health care"},{"id":56111808,"name":"Biological sciences/Microbiology"}],"tags":[],"updatedAt":"2025-12-01T15:59:10+00:00","versionOfRecord":{"articleIdentity":"rs-7414241","link":"https://doi.org/10.1038/s41598-025-30276-y","journal":{"identity":"scientific-reports","isVorOnly":false,"title":"Scientific Reports"},"publishedOn":"2025-11-28 15:56:54","publishedOnDateReadable":"November 28th, 2025"},"versionCreatedAt":"2025-10-13 14:17:23","video":"","vorDoi":"10.1038/s41598-025-30276-y","vorDoiUrl":"https://doi.org/10.1038/s41598-025-30276-y","workflowStages":[]},"version":"v1","identity":"rs-7414241","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7414241","identity":"rs-7414241","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}