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Abstract

Objective

This study aimed to isolate and identify bioactive secondary metabolites, including a new phenazine derivative, from a marine-derived Streptomyces sp. CNQ-031 belonging to the MAR4 clade, and to evaluate their antibacterial activity.

Methods

The marine-derived Streptomyces sp. was cultivated and extracted using ethyl acetate. The crude extracts were subjected to chromatographic separation to afford a new natural product, pontophenazine (1), along with known compounds, 1-methoxyphenazine (2), 5,7-dihydroxy-2-(1-methylpropyl)-4H-chromen-4-one (3), and 5,7-dihydroxy-2-isopropyl-4H-chromen-4-one (4). The chemical structure of 1 was elucidated through analysis of nuclear magnetic resonance (NMR) and mass spectrometry (MS) data, while the chemical structures of 2−4 were identified by comparing their NMR data with those of previously reported compounds. Antibacterial activity was assessed using the broth microdilution method.

Results

Four compounds were isolated, including a new phenazine derivative, pontophenazine (1) from a marine-derived Streptomyces sp. CNQ-031. This compound exhibited antibacterial activity against B. subtilis KCTC 1021, with a minimum inhibitory concentration (MIC) value of 64 μg/mL, showed significant biofilm inhibition towards A. tumefaciens SND195, and P. aeruginosa SNC165 (35.4 ± 1.1%, 19.7 ± 0.8%, respectively), and displayed weak inhibitoryactivity against M. luteus and S. aureus in the quorum-sensing assay.

Conclusion

The discovery of pontophenazine (1) expands the structural diversity of marine-derived phenazine derivatives and highlights the MAR4 Streptomyces clade as a valuable source of new antibacterial compounds.

Keywords

MAR 4 clade phenazine derivatives bioactivity chromone derivatives

Introduction

Natural products are a fundamental source of novel chemical diversity and an essential component of the pharmaceutical industry. Oceans, which cover 71% of the Earth's surface, contain 97% of the planet's water, and shelter nearly 87% of all animals and plants, are abundant sources of undiscovered organisms, including microorganisms. Among these, the exploration of rare actinomycetes within unexplored marine environments presents a promising avenue for discovering unprecedented chemical frameworks exhibiting a range of biological activities. These encompass anti-inflammatory, antibacterial, neuroprotective, antiparasitic, and anticancer properties.¹

Marine actinomycetes, particularly the Streptomyces lineage, have mainly originated from marine environments² and represent abundant sources for the discovery of interesting chemical scaffolds with bioactivity. The streptomycete clade MAR4, a distinct marine actinobacterial lineage, exhibits significant taxonomic diversity and excellent proficiency in producing diverse hybrid terpenoid skeletons such as polyketide-terpenoid molecules (particularly naphterpins D and E).³ Furthermore, MAR4 strains possess average five 2-amino-4-boronobutanoic acid (ABBA) prenyl-transferases per strain compared to other streptomycetes. Based on their substrate specificity, ABBA prenyl-transferases may be separated into two distinct group and are only present in secondary metabolism.^4,5 One family of ABBA, phenol/phenazine prenyl-transferases catalyses the prenylation of phenazines,⁶ naphtoquinones,⁷ benzodiazepines⁸ and hydroxybenzoates.⁹ The hybrid isoprenoids produced by MAR4 strains, including prenylated polyketides, pyrroles, and phenazines, are of considerable interest due to their potent biological activities.¹⁰ One of the strains belonging to this group, CNQ-031, isolated from marine sediment collected from La Jolla, California, has been reported to produce a phenazine derivative (2) and chromone derivatives (3, 4) with monoamine oxidase (MAO) activities.¹¹

Further investigation of the chemical components of this strain has resulted in the discovery of a new phenazine derivative, pontophenazine (1). This study describes the isolation and structure elucidation of a new compound, pontophenazine (1), as well as its biological activities, including previously reported compounds 2−4.

Experimental and Methods

General Experimental Procedure

The UV spectra were measured with a V-730 UV visible spectrophotometer (Jasco, Easton, MD, USA) using a path length of 1 cm. The infrared (IR) spectra were recorded on a Varian Scimitar Series (Palo Alto, CA, USA) using KBr disks. The NMR spectra were acquired with a Bruker NMR spectrometer (Bruker, Middlesex, MA, USA, 300 MHz for ¹H NMR) in MeOD-d₄ using a solvent signal as an internal reference (δ_H 3.31 ppm) and a JEOL NMR spectrometer (JEOL Ltd, Tokyo, Japan, 500 and 125 MHz for ¹H and ¹³C NMR, respectively) in DMSO-d₆ using a solvent signal as an internal reference (δ_H 2.50 ppm and δ_C 39.5 ppm). Low-resolution liquid chromatography/mass spectrometry (LC/MS) data were obtained on the Agilent Technologies (Santa Clara, CA, USA) 6120 quadrupole and Waters (Milford, MA, USA) Micromass ZQ LC/MS systems using a reversed-phase column (Phenomenex Luna C18(2), 100 Å, 50 mm × 4.6 mm, 5 μm) at a flow rate of 1.0 mL/min at the National Research Facilities and Equipment Center (NanoBio Energy Materials Center) at Ewha Womans University. High-resolution mass analysis was conducted by a JMS-700 mass spectrometer (JEOL Ltd, Tokyo, Japan) at Seoul National University. The fractions were purified by a Waters 616 quaternary HPLC pump and a Waters 996 photodiode array detector using a Phenomenex Luna C18(2) (100 Å, 250 mm × 10 mm, 5 μm) reversed-phase HPLC column.

Collection and Phylogenetic Analysis of Strain CNQ-031

The marine-derived actinomycetes strain, CNQ-031, was isolated from a sediment sampled off the coast of California. It was identified as the Streptomyces sp. MAR4 clade based on a 16S rRNA gene sequence analysis. The gene sequence data are available from GenBank (deposit number KC261620).

Fermentation and Extraction

The CNQ-031 strain was cultured in 40 L of a 2.5 L Ultra Yield Flask, each flask containing 1 L of medium SYP SW (10 g/L of soluble starch, 2 g/L of yeast, and 4 g/L of peptone dissolved in 1 L seawater), at 27 °C with shaking at 120 rpm for 7 days. The culture medium was extracted with EtOAc (40 L overall), and the EtOAc-soluble fraction was then evaporated in vacuo to yield 5.84 g of crude extract.

Isolation and Purification of Compounds

The organic extract of CNQ-031 (5.84 g) was fractionated with C-18 open column chromatography with a step gradient from 20% to 100% MeOH in DW to obtain nine fractions. The third fraction was further subjected to reversed-phase HPLC with 39% aqueous acetonitrile (Phenomenex Luna C18(2), 250 × 100 mm, 2.0 mL/min, 5 μm, 100 Å) to obtain 5.0 mg of pontophenazine (1) at t_R = 16 min and 5,7-dihydroxy-2-isopropyl-4H-chromen-4-one (4, t_R = 39 min, 19.8 mg). To isolate 1-methoxyphenazine (2, t_R = 26 min, 6.0 mg), the fourth fraction was chromatographed by elution with 50% acetonitrile. The fifth fraction was purified by 47% acetonitrile to obtain 5,7-dihydroxy-2-(1-methylpropyl)-4H-chromen-4-one (3, t_R = 33 min, 11.4 mg).

Pontophenazine (1): deep yellow amorphous solid; UV (CH₃OH) λ_max (log ε) 202 (3.4), 220 (3.4), 240 (3.3), 264 (3.2), 400 (2.9) nm; IR (KBr) ν_max 2957, 2925, 2351, 1728, 1454 cm⁻¹, ¹H-NMR (DMSO-d₆, 500 MHz) δ_H 6.99 (1H, dd, J = 7.8, 1.4 Hz, H-3), 6.75 (1H, t, J = 7.8 Hz, H-4), 6.55 (1H, dd, J = 7.8, 1.4 Hz, H-5), 3.00 (3H, s, H-8); ¹³C-NMR (DMSO-d₆, 125 MHz) δ_C 168.7 (C-1), 143.5 (C-7), 130.9 (C-6), 123.1 (C-3), 120.8 (C-4), 118.6 (C-2), 114.4 (C-5), 39.1 (C-8) see Table S1; HR-FAB-MS [M]⁺ m/z 298.0955.

Antibacterial Activity Testing (MIC Assay)

Antibacterial activity was evaluated against a panel of Gram-positive and Gram-negative bacteria. The Gram-positive bacteria used were Bacillus subtilis KCTC1021, Staphylococcus aureus KCTC1927, Kocuria rhizophila KCTC1915, and Micrococcus luteus SC0560. The Gram-negative bacteria included Escherichia coli KCTC2441, Klebsiella pneumoniae KCTC2690, Salmonella typhimurium KCTC2515, Pseudomonas fluorescens SND204, Pseudomonas aeruginosa SNC165, and Agrobacterium tumefaciens SND195.

All bacterial strains were grown overnight in the appropriate media (Mueller-Hinton Broth for the KCTC strains, tryptic soy broth for S. aureus and M. luteus, Kings broth for P. aeruginosa and P. fluorescens, and marine broth for A. tumefaciens). Cultures were adjusted to a 0.5 McFarland standard (1.5 × 10⁸ CFU/mL).

The test compounds and positive controls (vancomycin, ampicillin, kanamycin, and rifampin) were dissolved in DMSO at a concentration of 256 μg/mL, and DMSO was used as the negative control. Each compound (100 μL) was added to the first well of a sterile 96-well plate containing 50 μL of the respective growth medium, followed by two-fold serial dilutions to obtain final concentrations of 128, 64, 32, 16, 8, 4, 2, 1, 0.5, and 0.25 μg/mL.

Subsequently, 50 μL of the adjusted bacterial inoculum was added to each well, resulting in a final inoculum density of 5.0 × 10⁵ CFU/mL. Plates were incubated at 37 °C for 18–24 h. MIC values were determined by visual examination of bacterial growth inhibition; clear wells indicated growth inhibition, whereas turbid wells indicated bacterial growth. All experiments were performed in triplicate, and the results were reproducible within experimental error; therefore, no statistical analysis was applied.^12,13

Biofilm Inhibition Activity

The biofilm inhibition activity was tested using P. fluorescens SND204, P. aeruginosa SNC165, A. tumefaciens SND195, S. aureus KCTC1927, and M. luteus SC0560 as the biosensors. After cultivating the bacteria in the same way as in the quorum sensing test, compounds were inoculated (256 µg/mL, 200 μL) in each culture and cultivated for 72 h at 37 °C. Briefly, 2 mL of media and 50 μL of the tested compound, positive controls (rifampin and kanamycin) and negative control (DMSO) were added into individual 10 mL test tubes. Each test tube was then inoculated with 4 μL of appropriately adjusted bacterial cultures and incubated at 37 °C for 72 h without shaking. After completing the incubation, the cultures were gently discarded using a 1 mL pipette, ensuring caution to avoid damaging the biofilm formed on the tube walls. Subsequently, the tubes underwent three washes with sterile distilled water. The bacteria constituting the biofilm were stained by adding 2.5 mL of 0.1% crystal violet for 1 h without shaking. The crystal violet dye was discarded, and the tubes were washed with sterile distilled water to eliminate any unbound dye. The tubes were then dried, and the biofilm was eluted using 2 mL of 100% ethanol. Thorough dissolution of the dye was ensured by shaking the tubes. Finally, the optical density (OD_590nm) of the eluted samples was measured using a spectrophotometer.¹⁴ All experiments were performed in triplicate, and the results were reproducible within experimental error; therefore, no statistical analysis was applied.

Result

Pontophenazine (1) was isolated as a deep yellow amorphous solid, and its molecular formula was deduced as C₁₆H₁₄N₂O₄ based on high-resolution-fast atom bombardment-mass spectrometry (HR-FAB-MS) data [M]⁺ at m/z 298.0955 (calculated for [C₁₆H₁₄N₂O₄]⁺, 298.0948), coupled with NMR data analysis. The IR spectrum of 1 showed absorption bands for the carboxyl group at 1728cm⁻¹. The ¹H NMR spectroscopic data of 1 indicated the presence of three aromatic protons at δ_H 6.99 (1H, dd, J = 7.8, 1.4 Hz, H-3), 6.75 (1H, t, J = 7.8 Hz, H-4), and 6.55 (1H, dd, J = 7.8, 1.4 Hz, H-5), and an N-methyl group at δ_H 3.00 (3H, s, H-8). The ¹³C NMR data and heteronuclear single quantum coherence (HSQC) spectrum of 1 revealed eight carbons including a carbonyl carbon at δ_C 168.7 (C-1), three quaternary carbons at δ_C 143.5 (C-6), 139.0 (C-7), and 118.6 (C-2), three methine carbons at δ_C 123.1 (C-3), 120.8 (C-4), and 114.4 (C-5), and an N-methyl carbon at δ_C 39.1 (C-8).

Analysis of 2D NMR spectroscopic data allowed the structural assignment of 1 (Table S1). The 12,3-tri-substituted benzene ring system was deduced from the observation of ¹H-¹H homonuclear COSY correlations of H-3 (dd, J = 7.8, 1.4 Hz) and H-5 (dd, J = 7.8, 1.4 Hz) to H-4 (t, J = 7.8 Hz). Further, three-bond ¹H-¹³C heteronuclear multiple bond correlations (HMBC) of H-4 and N-CH₃ to C-6 suggested that the N-CH₃ group should be attached at C-6. In a similar fashion, the observation of HMBC correlations from N-CH₃ to C-6 and C-7′, from H-3 to C-1, C-5, and C-7, from H-4 to C-2 and C-6, and from H-5 to C-3 and C-7 allowed the phenazine moiety to be defined. In addition, the three-bond HMBC correlation from H-3 to C-1 revealed the attachment of a carboxylic acid group to the phenazine moiety at C-1 in pontophenazine (1).

Based on the molecular formula and the number of carbon signals observed in the ¹³C NMR spectrum, the chemical structure of 1 was designated as a symmetrical structure, as shown in Figure 1a.

Figure 1.

The chemical structures of 1–4.

Compound 2 was identified as 1-methoxy phenazine after comparing its NMR data to that of the previously reported literature,¹⁵ while compounds 3 and 4 were identified as hyperfaberol F (5,7-dihydroxy-2-(1-methylpropyl)-4H-chromen-4-one) and 7-dihydroxy-2-isopropyl-4H-chromen-4-one, respectively, by comparing the MS and NMR spectroscopic data with those in the literature.¹⁶

Pontophenazine (1) and 1-methoxy phenazine (2) belong to the well-known synthetic and natural phenazine groups, which have been extensively studied for their potential use in various fields, such as anticancer and cystic fibrosis agents. Reported phenazine compounds, such as phenazine-1-carboxylic acid (PCA), pyocyanin, 5-methyl phenazine-1-carboxylic acid, phenaziterpene A, and phenaziterpene B are mostly isolated from terrestrial and marine microorganisms. PCA, also known as tubermycin B, is prevalent across a wide range of microorganisms and has shown anti-fungal and anticancer activities. Phenazines with a symmetric structure such as phenazine-1,6-dicarboxylate, 1,6-dihydroxyphenazine-5,10-dioxide, and 1,6-phenazinedimethanol have also been reported.^17–18 Compound 2 was previously isolated from Nocardiopsis dassonvillei and was found to exhibit inhibitory activities against the K562, A549, and MCF-7 cell lines, H1N1 virus, MAO-A, and MAO-B.¹⁹ Furthermore, pontophenazine (1) was the first compound to be isolated with a symmetric structure containing two N-methyl groups.

Compounds 3 and 4 are categorized as chromones, also referred to as 4H-chromen-4-ones, 4H-1-benzopyran-4-ones, or benzo-γ-pyrones. These compounds belong to a class of heterocyclic compounds containing oxygen, characterized by a benzopyran derivative with a substituted keto group on the pyran ring.²⁰ Chromones are widely studied due to their advantageous drug-like properties and versatile binding properties, making them privileged scaffolds in medicinal chemistry. These compounds exhibit diverse biological activities including anti-inflammatory, antioxidant, cardioprotective, and hepaprotective effects. Compounds 3 and 4 have been reported to display inhibitory activities against MAO-A and MAO-B.¹¹ In addition, compound 3, isolated from Hypericum sp., has been reported as both a synthetic and natural product.^21,22 Meanwhile, compound 4 was first produced from flowering plants of the genus Helichrysum²³ and was found to exhibit cytotoxicity against HepG2.²⁴ This is the first isolation of compound 4 from bacteria.

To explore the antibacterial properties of the isolated compounds, samples 1–4 were evaluated against a panel of Gram-positive and Gram-negative bacteria using the MIC assay. As shown in Table 1, compounds 1 and 4 exhibited weak antibacterial activity toward Bacillus subtilis KCTC1021, each showing an MIC value of 64 μg/mL. Compound 3 also demonstrated weak inhibitory activity, with an MIC value of 64 μg/mL against both B. subtilis KCTC1021 and Staphylococcus aureus KCTC1927. Additionally, compound 1 showed moderate inhibitory activity toward Micrococcus luteus SC0560 with an MIC of 32 μg/mL, whereas compound 3 inhibited the same strain at 64 μg/mL. Compound 3 also exhibited significant activity against Agrobacterium tumefaciens SND195 with an MIC value of 16 μg/mL. Overall, the tested compounds displayed selective and generally weak antibacterial activities, with the lowest MIC value observed for compound 3 against A. tumefaciens SND195 (MIC = 16 μg/mL).

Table 1.

Antibacterial Activities of Compounds 1−4.^a

Compound	MIC (μg/mL)
	Gram (+) Bacteria				Gram (–) Bacteria
	B. subtilis KCTC1021	K. rhizophila KCTC1915	S. aureus KCTC1927	M.luteusSC0560	E. coli KCTC2441	S. typhimurium KCTC2515	K. pneumonia KCTC2690	P. aeruginosaSNC165	P.fluorescensSND204	A.tumefaciensSND195
1	64	>128	>128	32	>128	>128	>128	>128	>128	>128
2	>128	>128	>128	>128	>128	>128	>128	>128	>128	>128
3	64	>128	64	64	>128	>128	>128	>128	>128	16
4	64	>128	>128	>128	>128	>128	>128	>128	>128	>128
Ampicillin	0.25	0.25	0.25	-	4	2	>128	-	-	-
Vancomycin	0.25	0.5	0.5	-	>128	>128	>128	-	-	-
Kanamycin	-	-	-	64	-	-	-	>128	8	>128
Rifampin	-	-	-	0.25	-	-	-	32	16	2
DMSO	>128	>128	>128	>128	>128	>128	>128	>128	>128	>128

Each sample was tested in triplicate, and the experiment was repeated three times.

In the antibiofilm assay, compound 3 exhibited notable biofilm inhibition towards A. tumefaciens SND195, S. aureus KCTC1927, and P. aeruginosa SNC165 (70.3 ± 0.5%, 47.4 ± 1.7%, and 27.3 ± 3.4%, respectively). Compound 1 showed moderate inhibitory effects on biofilm formation of A. tumefaciens SND195 and P. aeruginosa SNC165 (35.4 ± 1.1% and 19.7 ± 0.8%, respectively). Additionally, compound 4 exhibited weak biofilm inhibition against P. aeruginosa SNC165 (10.9 ± 1.5%), whereas compound 2 showed no observable inhibition (Figure 2).

Figure 2.

Biofilm inhibition activities of 1-4. Percentage of biofilm inhibition refers to the percentage of biofilm that was inhibited in the presence of the compounds, when compared to biofilm formation with only DMSO. Shown are the average results of three replicates and error bars represent the standard error of the mean (SEM).

Discussion

The biological properties of phenazines are determined by the positions and types of functional groups present in their molecules.²⁵ Phenazines and their derivatives function as electron shuttles, regulating the redox state of cells and influencing downstream gene expression associated with biofilm formation and bacterial survival.²⁶ Based on previous study,¹⁸ the position of the substituents in phenazine derivatives greatly influences their antibacterial activity. Moreover, according to the QSAR analysis of the antibacterial mechanism of phenazines conducted by Udumula et al²⁷ 2017, it is anticipated that compounds with higher water solubility and lower EHOMO energies may have more potent antibacterial action. Compound 1 exhibited antibacterial and biofilm inhibition activity, whereas compound 2 displayed no biological activity. Moreover, phencomycin, PCA, and phenazine-1,6-dicarboxylate which are phenazine derivatives containing carbonyl carbons such as carboxylic acid and ester groups, have been reported to exhibit notable antibiofilm and antibacterial activities against four bacterial strains (E. coli, S. aureus, P. aeruginosa, and B. subtilis) with minor cytotoxicity against four cell lines (WI38, HCT116, HepG-2 and MCF-7).²⁸ 5-methyl phenazine-1-carboxylic acid, a phenazine derivatives containing N-methyl and carboxylic acid, showed selective cytotoxicity against A549 and MDA MB-231 cell lines. Therefore, these findings suggested that carbonyl groups and N-methyl substituents play important roles in exhibiting the biological activities.

Biofilm formation is a crucial virulence and persistence strategy for many pathogenic bacteria, as cells embedded in an extracellular polymeric substance (EPS) matrix show increased tolerance to antibiotics and host immune responses compared to their planktonic counterparts.^29,30 This enhanced tolerance contributes to chronic and relapsing infections and is recognized as an important factor in antimicrobial resistance.^31,32 In this study, compound 3 demonstrated the most significant antibiofilm effect, especially against A. tumefaciens SND195, S. aureus KCTC1927, and P. aeruginosa SNC165, indicating its ability to interfere with biofilm development in both Gram-positive and Gram-negative bacteria. A. tumefaciens and P. aeruginosa are well known for forming strong biofilms on biotic and abiotic surfaces, which promotes colonization and persistence in environmental and clinical settings, respectively.^33,34 The moderate antibiofilm activity observed for compound 1 and the weak or negligible effects of compounds 4 and 2 suggest a structure–activity relationship that merits further study. Like other natural or semi-synthetic small molecules reported to inhibit biofilm formation in S. aureus, P. aeruginosa, and plant-associated pathogens,^31,35,36 the antibiofilm profile of compound 3 underscores its potential as a lead scaffold for developing new agents targeting biofilm-related infections or biofilm-driven colonization in agrobiotechnological applications.

Although, pontophenazine (1) was successfully identified as a new phenazine analog with moderate antibacterial and quorum sensing inhibitory activities, this study remains limited to in vitro evaluations. The mechanism of action, cytotoxicity, and biosynthetic origin of the compound were not investigated. Future studies focusing on biosynthetic gene cluster analysis and structure–activity relationship optimization are necessary to further explore its pharmacological potential.

Conclusions

A new natural product, pontophenazine (1), was isolated from the marine-derived Streptomyces sp. MAR4 CNQ-031, along with three known compounds, 1-methoxyphenazine (2), 5,7-dihydroxy-2-(1-methylpropyl)-4H-chromen-4-one (3), and 5,7-dihydroxy-2-isopropyl-4H-chromen-4-one (4). Based on the antibacterial and antibiofilm evaluations, compound 3 demonstrated the strongest overall biological activity, exhibiting the lowest MIC value (16 μg/mL) and the highest levels of biofilm inhibition across multiple bacterial strains. Compound 1 showed moderate but selective antibacterial and antibiofilm effects, while compounds 2 and 4 displayed minimal to weak activity. These findings indicate that compound 3 possesses the most promising antimicrobial potential among the tested isolates.

Supplemental Material

sj-docx-1-npx-10.1177_1934578X251405973 - Supplemental material for Pontophenazine, a new Phenazine Analog from Marine-Derived Streptomyces sp. CNQ-031

Supplemental material, sj-docx-1-npx-10.1177_1934578X251405973 for Pontophenazine, a new Phenazine Analog from Marine-Derived Streptomyces sp. CNQ-031 by Chaeyoung Lee, Prima F Hillman, Ji Young Lee, Dong-Chan Oh and Songyi Lee, William Fenical, Sang-Jip Nam in Natural Product Communications

Footnotes

ORCID iDs

Chaeyoung Lee

Prima F Hillman

Sang-Jip Nam

Ethical Approval Statement

This article does not involve studies with human participants or animals. Ethical approval was therefore not required for this research.

Statement of Informed Consent

There are no human subjects in this article, and informed consent is not applicable.

Author Contributions

Compound isolation, structure elucidation, activity test and manuscript writing, C.L., P.F.H.; activity test J.Y.L.; writing—review and editing, C.L., P.F.H., D.-C.O.; supervision and conceptualization, S.L., W.F., S.-J.N. All authors have read and agreed to the published version of the manuscript.

Funding

The authors disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: This research was supported by Korea Basic Science Institute (National Research Facilities and Equipment Center) grant funded by the Ministry of Education (2020R 1A 6C 101B194 to S.-J.N.); the National Research Foundation of Korea (NRF) grant funded by the Korean government (MSIT; No. NRF-2022R 1A 2C 1011848 to S.-J.N.). This research was supported by the Global Learning & Academic Research Institution for Master's·PhD students, and Postdocs (LAMP) Program of the National Research Foundation of Korea (NRF) grant funded by the Ministry of Education (No. RS-2023-00301702). Isolation of the bacterium was a result of the financial support from the US National Cancer Institute (grant CA R37044848 to W.F.).

Declaration of Conflicting Interests

The authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Data Availability Interests

Data available on request from the corresponding author.

Reporting Guidelines Statement

Reporting guidelines such as CONSORT or ARRIVE do not directly apply to this in vitro study; however, we have followed best practices for transparent and reproducible reporting.

Supplemental Material

Supplemental material for this article is available online.

Note

This article does not contain any studies on human or animal subjects, and the statement of human and animal rights is not applicable.

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Supplementary Material

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Pontophenazine,a new Phenazine Analog from Marine-Derived Streptomyces sp. CNQ-031

Abstract

Objective

Methods

Results

Conclusion

Keywords

Introduction

Experimental and Methods

General Experimental Procedure

Collection and Phylogenetic Analysis of Strain CNQ-031

Fermentation and Extraction

Isolation and Purification of Compounds

Antibacterial Activity Testing (MIC Assay)

Biofilm Inhibition Activity

Result

Discussion

Conclusions

Supplemental Material

sj-docx-1-npx-10.1177_1934578X251405973 - Supplemental material for Pontophenazine, a new Phenazine Analog from Marine-Derived Streptomyces sp. CNQ-031

Footnotes

ORCID iDs

Ethical Approval Statement

Statement of Informed Consent

Author Contributions

Funding

Declaration of Conflicting Interests

Data Availability Interests

Reporting Guidelines Statement

Supplemental Material

Note

References

Supplementary Material