A quantitative analytical method for valienone and its application in the evaluation of valienone production by a breakthrough microbial process

A Validamycin Shunt Pathway for Valienamine Synthesis in Engineered Streptomyces hygroscopicus 5008

“…The levels of the varB transcript were monitored using quantitative real-time PCR (qPCR) every 24 h, and the production of valienamine was quantified using LC-TQMS and confirmed by comparing the retention time and the MRM fragmentation of the standard (Figure S6). Moreover, valienone and validamycin A were detected using a precolumn DNPH-derivative HPLC 34 and HPLC analyses, respectively.…”

“…20 The precolumn derivative HPLC assay for valienone was performed using an Eclipse XDB-C18 column (5 μm, 4.6 × 150 mm) at 30 °C with 50% acetonitrile, and the absorbance was monitored at 380 nm after derivatization with 2,4-dinitrophenylhydrazine in a phosphate solution at 37 °C for 45 min. 34 TQMS for valienamine was conducted on Agilent QQQ MS 6470 using an Eclipse XDB-C18 (5 μm, 4.6 × 150 mm) column; the 22% acetonitrile eluent at 30 °C was directed to the MS with an ESI source gas temperature of 300 °C, a gas flow rate of 5 L/min, and a nebulizer pressure of 45 psi. The quantification of small molecular compounds was based on the integrated peak area of the MRM chromatograms.…”

“…The HPLC analysis of validamycin A was performed using an Eclipse XDB-C18 column (5 μm, 4.6 × 150 mm) with 3% acetonitrile at 30 °C and monitored at 210 nm using a diode array detector . The precolumn derivative HPLC assay for valienone was performed using an Eclipse XDB-C18 column (5 μm, 4.6 × 150 mm) at 30 °C with 50% acetonitrile, and the absorbance was monitored at 380 nm after derivatization with 2,4-dinitrophenylhydrazine in a phosphate solution at 37 °C for 45 min …”