Discovery of Undescribed Clerodane Diterpenoids with Antimicrobial Activity Isolated from the Roots of Solidago gigantea Ait

Three previously undescribed clerodane diterpenoids, including two <i>cis</i>-clerodanes, solidagolactone IX (<b>1</b>) and solidagoic acid K (<b>2</b>), and one <i>trans</i>-clerodane, solidagodiol (<b>3</b>), along with two known <i>cis</i>-clerodane diterpenoids, (-)-(5<i>R</i>,8<i>R</i>,9<i>R</i>,10<i>S</i>)-15,16-epoxy<i>-ent</i>-<i>neo</i>-cleroda-3,13,14-trien-18-ol (<b>4</b>) and solidagoic acid J (<b>5</b>), were isolated and comprehensively characterized from the ethanolic and ethyl acetate root extract of <i>Solidago gigantea</i> Ait. (giant goldenrod). Compound <b>4</b> has previously been reported from the roots of this species, whereas compound <b>5</b> was identified from the leaves of <i>S. gigantea</i> but not from the roots. The bioassay-guided isolation involved thin-layer chromatography-direct bioautography (TLC-DB) with a <i>Bacillus subtilis</i> antibacterial assay, preparative flash column chromatography, and TLC-mass spectrometry (MS). The chemical structures of the isolated compounds (<b>1</b>-<b>5</b>) were elucidated through extensive in-depth spectroscopic and spectrometric analyses, including one- and two-dimensional nuclear magnetic resonance (NMR) spectroscopy, high-resolution tandem mass spectrometry (HRMS/MS), and attenuated total reflectance Fourier-transform infrared (ATR-FTIR) spectroscopy. Their antimicrobial activities were evaluated using <i>in vitro</i> microdilution assays against <i>B. subtilis</i> and different plant pathogens. Compound <b>3</b> was the most active against the tested Gram-positive strains, exerting particularly potent effects against <i>Clavibacter michiganensis</i> with a minimal inhibitory concentration (MIC) value of 5.1 µM as well as <i>B. subtilis</i> and <i>Curtobacterium flaccumfaciens</i> pv. <i>flaccumfaciens</i> (MIC 21 µM for both). Compound <b>4</b> also strongly inhibited the growth of <i>C. michiganensis</i> (MIC 6.3 µM). Compounds <b>2</b>, <b>4</b>, and <b>5</b> displayed moderate to weak activity against <i>B. subtilis</i> and <i>C. flaccumfaciens</i> pv. <i>flaccumfaciens</i> with MIC values ranging from 100 to 402 µM. <i>Rhodococcus fascians</i> bacteria were moderately inhibited by compounds <b>3</b> (MIC 41 µM) and <b>4</b> (MIC 201 µM). Bactericidal activity was observed for compound <b>3</b> against <i>C. michiganensis</i> with a minimal bactericidal concentration (MBC) value of 83 µM. Compounds <b>2</b> and <b>3</b> demonstrated weak antifungal activity against <i>Fusarium graminearum</i>. Our findings underscore the value of bioassay-guided approaches in discovering previously undescribed bioactive compounds.