An in vitro study of the antimicrobial effects of indigo naturalis prepared from Strobilanthes formosanus Moore

Yin-Ru Chiang, Ann Li, Yann-Lii Leu, Jia-You Fang, Yin-Ku Lin, Yin-Ru Chiang, Ann Li, Yann-Lii Leu, Jia-You Fang, Yin-Ku Lin

Abstract

Indigo naturalis is effective in treating nail psoriasis coexisting with microorganism infections. This study examines the antimicrobial effects of indigo naturalis prepared from Strobilanthes formosanus Moore. Eight bacterial and seven fungal strains were assayed using the agar diffusion method to examine the effects of indigo naturalis and its bioactive compounds. The bioactive compounds of indigo naturalis were purified sequentially using GFC, TLC, and HPLC. Their structures were identified using mass spectrometry and NMR spectroscopy. UPLC-MS/MS was applied to compare the metabolome profiles of indigo naturalis ethyl-acetate (EA) extract and its source plant, Strobilanthes formosanus Moore. The results of in vitro antimicrobial assays showed that indigo naturalis EA-extract significantly (≥1 mg/disc) inhibits Gram-positive bacteria (Staphylococcus aureus, S. epidermis and methicillin-resistant S. aureus (MRSA)) and mildly inhibits non-dermatophytic onychomycosis pathogens (Aspergillus fumigates and Candida albicans), but has little effect on dermatophyes. Isatin and tryptanthrin were identified as the bioactive compounds of indigo naturalis using S. aureus and S. epidermis as the bioassay model. Both bioactive ingredients had no effect on all tested fungi. In summary, indigo naturalis prepared from Strobilanthes formosanus Moore exhibits antimicrobial effects on Staphylococcus and non-dermatophytic onychomycosis pathogens. Tryptanthrin and isatin may be its major bioactive ingredients against Staphylococcus and the inhibitory effect on MRSA may be due to other unidentified ingredients.

Figures

Figure 1
Figure 1
APCI-MS and 1H-NMR spectra of the anti-Staphylococcus compounds (IN-1 and IN-2) purified from the EA-extract of indigo naturalis; (A) APCI-mass spectrum of the compound IN-1; (B) APCI-mass spectrum of the compound IN-2; (C) 1H-NMR spectrum of the compound IN-1 (400 MHz, acetone-d6); (D) 1H-NMR spectrum of the compound IN-2 (400 MHz, acetone-d6).
Figure 2
Figure 2
UPLC-APCI-MS analysis of the standards (isatin and tryptanthrin) and EA-extracts of indigo naturalis and Strobilanthes formosanus Moore. (A) APCI-mass spectra of isatin (A-1) and tryptanthrin (A-2) purchased from Sigma-Aldrich Chemicals; (B) UPLC chromatogram of the mixture (molar ratio = 1:1) of isatin and tryptanthrin purchased from Sigma-Aldrich Chemical; (C) UPLC total ion chromatogram of the EA-extract of indigo naturalis powder (50 µg); (D) UPLC total ion chromatogram of the EA-extract of the dry leaves from S. formosanus (50 µg).
Figure 3
Figure 3
HPLC quantification of the anti-Staphylococcus compounds (isatin and tryptanthrin) in the EA-extracts of the indigo naturalis powder and the dry leaves from Strobilanthes formosanus Moore.

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