Vitroprocines, new antibiotics against Acinetobacter baumannii, discovered from marine Vibrio sp. QWI-06 using mass-spectrometry-based metabolomics approach

A robust and convenient research strategy integrating state-of-the-art analytical techniques is needed to efficiently discover novel compounds from marine microbial resources. In this study, we identified a series of amino-polyketide derivatives, vitroprocines A-J, from the marine bacterium Vibrio sp. QWI-06 by an integrated approach using imaging mass spectroscopy and molecular networking, as well as conventional bioactivity-guided fractionation and isolation. The structure-activity relationship of vitroprocines against Acinetobacter baumannii is proposed. In addition, feeding experiments with 13C-labeled precursors indicated that a pyridoxal 5′-phosphate-dependent mechanism is involved in the biosynthesis of vitroprocines. Elucidation of amino-polyketide derivatives from a species of marine bacteria for the first time demonstrates the potential of this integrated metabolomics approach to uncover marine bacterial biodiversity.


Figure S66
EIMS spectrum and fragmentation of the dimethyl disulfide derivative of 8 48

Figure S67
UV spectrum of 9 49

Figure S71
UV spectrum of 10 51

Figure S75
EIMS spectrum and fragmentation of the dimethyl disulfide derivative of 10 53

Figure S78
13 C NMR spectra of 1 and 1a 56 Experimental Section Instrumentation. Optical rotations were measured on a JASCO P1020 digital polarimeter at 25 °C. UV spectra were recorded on a JASCO V-650 spectrophotometer. IR spectra were obtained on a JASCO FT/IR-4100 infrared spectrophotometer. NMR spectra were recorded with Bruker 500 AVII and Varian Unity 400 FT NMR spectrometers. HPLC-ESIMS data were processed using an AmaZon SL ion trap mass spectrometer (Bruker Daltonics, Bremen, Germany) and UPLC-HR-ESIMS and MS/MS data were acquired on a Thermo Orbitrap Elite system and a Thermo TSQ Quantum Access MAX Triple Quadrupole system. IMS data were acquired in a Bruker autoflex TM speed MALDI-TOF/TOF system. Extracts were chromatographed on Sephadex LH-20 (Amersham Biosciences) and purified using a semi-preparative RP HPLC column (Supelco Discovery C18, 250 x 10 mm, 5 μm). HPLC was performed on a Hitachi L-2130 apparatus equipped with a Hitachi L-2420 UV-VIS detector. Biosynthetic studies. 1-13 C-sodium acetate and 13 C 9 -tyrosine were used (99% of 13 C, Cambridge Isotope Laboratories, Isotec, and Aldrich) for the isotope labeling experiments. Vibrio sp. QWI-06 was cultivated in Difco marine agar. For each experiment, 0.1 mM of 1-13 C-sodium acetate and 13 C 9 -tyrosine were respectively added in the sterile agar medium for 50 Petri dish plates (90 × 15 mm). Following the inoculum, all the cultures were grown for three days at 30 °C. The isolation of labeled vitroprocine A (1) was carried out according to the aforementioned procedure and the compound was subjected to NMR and LC-HR-ESIMS/MS analysis (Supporting information Figures S77 and S78). The yields of 1-13 C-sodium acetate labeled vitroprocine A (1a) were approximately 0.2 mg/plate.         Figure Figure Figure S77. HR-ESIMS/MS spectrum and fragmentation of 1 from Vibrio sp. QWI-06 feed with 13 C 9 -tyrosine. Figure S78. 13 C NMR spectra of 1 and 1a. 1a was isolated from Vibrio sp. QWI-06 feed with 1-13 C-sodium acetate. The enhanced carbon signals of 1a were marked in red.