INTRODUCTION

Antimicrobial resistance is one of the greatest global public health challenges, accounting for an estimated 4.95 million deaths annually, with projections indicating that mortality could reach 10 million deaths per year by 2050 if effective interventions are not implemented [1–5]. Neisseria gonorrhoeae and Candida albicans are priority pathogens identified by the World Health Organization because of their increasing resistance to conventional antibiotics and antifungal agents, which has intensified the global burden of sexually transmitted infections and vulvovaginal candidiasis, affecting approximately 87 million and 138 million individuals annually, respectively [6–8]. N. gonorrhoeae has developed resistance to penicillin, tetracycline, fluoroquinolones, and third-generation cephalosporins, whereas C. albicans has acquired resistance to azoles and echinocandins through multiple mechanisms, including target-site mutations, efflux pump overexpression, and biofilm formation [9–12]. Coinfections involving these pathogens frequently occur in the reproductive tract, manifesting clinically as urethritis, cervicitis, and vulvovaginitis, and often require combined antibacterial and antifungal therapy. However, such treatment may increase the risk of adverse drug reactions, drug interactions, treatment costs, and the continued emergence of antimicrobial resistance [6]. Consequently, there is increasing interest in identifying plant-derived therapeutic agents possessing multiple mechanisms of action that may serve as effective alternatives or complementary therapies to conventional antimicrobial agents [13, 14].

Oxidative stress also plays an important role in the pathogenesis of genital infections. Excessive production of reactive oxygen species (ROS) promotes tissue damage, disrupts epithelial integrity, and weakens local immune defenses, thereby facilitating pathogen persistence and disease progression. Plant-derived antioxidant compounds may provide dual therapeutic benefits by directly inhibiting microbial growth while simultaneously reducing oxidative tissue injury [15–21]. Therefore, evaluating both the antimicrobial and antioxidant activities of medicinal plants is essential for developing multifunctional therapeutic agents that eliminate pathogens while preserving host tissue integrity. Indonesia possesses exceptional plant biodiversity, with more than 30,000 plant species, approximately 7,000 of which are recognized for their medicinal properties and have traditionally been used to treat various infectious diseases [22, 23]. Among these medicinal plants, Coleus scutellarioides (L.) Benth. (synonyms: Coleus blumei, Plectranthus scutellarioides, and Solenostemon scutellarioides), commonly known as Miana or jawer kotok, belongs to the family Lamiaceae and has long been used in traditional medicine to treat cough, fever, diarrhea, skin infections, and respiratory disorders [24–27]. Phytochemical investigations have demonstrated that Miana leaves contain flavonoids, terpenoids, alkaloids, saponins, tannins, and essential oils exhibiting antibacterial, antifungal, anti-inflammatory, and antioxidant activities [15–21]. Tarigan et al. [17] reported that methanolic extracts of Miana leaves inhibited the growth of Staphylococcus aureus, whereas Bismelah et al. [28] demonstrated antibacterial activity against periodontal bacteria, reporting a minimum inhibitory concentration (MIC) of 1.56 mg/mL and a minimum fungicidal concentration (MFC) of 3.13 mg/mL.

Despite these promising findings, several important knowledge gaps remain. Previous studies have largely relied on conventional phytochemical techniques, such as thin-layer chromatography and spectrophotometry, which provide only limited information regarding the comprehensive metabolite composition of Miana. Furthermore, most investigations have focused primarily on gram-positive bacteria, oral pathogens, or dermatophytes and have not evaluated clinically important sexually transmitted pathogens, such as N. gonorrhoeae and C. albicans, using standardized Clinical and Laboratory Standards Institute (CLSI)-based antimicrobial susceptibility testing methods. Although the antioxidant properties of Miana leaves have been independently reported by Ślusarczyk et al. [15] and Pakadang et al. [20], these activities have not been systematically investigated alongside antimicrobial activity within a single, integrated experimental framework. Moreover, although previous studies have demonstrated in vivo antifungal effects against C. albicans in murine vulvovaginal infection models and antibacterial activity against oral and gram-positive pathogens, no published study has comprehensively combined untargeted metabolomic profiling with standardized in vitro MIC and MFC determination against both N. gonorrhoeae and C. albicans. Consequently, the relationships between the metabolomic composition of Miana extracts and their dual antimicrobial and antioxidant activities remain poorly understood, thereby limiting the scientific evidence needed to develop standardized phytopharmaceutical products.

Therefore, this study aimed to comprehensively characterize the secondary metabolite profile of ethanol extracts of Miana leaves using untargeted liquid chromatography–high-resolution mass spectrometry (LC–HRMS). In addition, the antimicrobial activities of the extracts against N. gonorrhoeae and C. albicans were evaluated by determining the MIC and MFC values using the standardized CLSI broth microdilution method, while antioxidant activity was assessed using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging assays. Untargeted LC–HRMS enables simultaneous identification of diverse secondary metabolites with high mass accuracy (<5 ppm) and compound-level annotation confidence in accordance with the Metabolomics Standards Initiative guidelines, thereby providing a comprehensive phytochemical profile that supports extract standardization and structure–activity relationship analyses [29]. The selection of N. gonorrhoeae and C. albicans as test microorganisms is clinically relevant because both pathogens are major causes of genital infections, frequently coinfect, and exhibit increasing antimicrobial resistance, often necessitating combination therapy [30, 31]. The findings of this study are expected to provide robust scientific evidence supporting the dual antimicrobial and antioxidant potential of Miana leaf extracts, identify bioactive metabolites associated with these biological activities, and establish a comprehensive phytochemical foundation for the future development of standardized phytopharmaceutical agents for the management of gonorrhea, candidiasis, and other genital infections.