Avance neo 400 mhz spectrometer

Cultures of 20 mL X. autotrophicus cells were grown to OD₆₀₀ = 2.0 over 72 h, then centrifuged at 4,000 rpm (10 min at 4 °C). For the whole-cell solution NMR spectra, about 100 mg of wet cell pellet was resuspended in 200 μL fructose-supplemented liquid Schuster minimal media. Fresh cell spectra were obtained from 100 mg of a fresh cell pellet, while frozen-cell spectra were obtained from 100 mg of a cell pellet that had been stored at −20 °C for 24 h to mimic the SSNMR sample condition prior to resuspension. After respension, both samples appear homogeneous, opaque and yellow. Cell extract spectra were obtained from the soluble fraction of fresh cell pellet or from the soluble fraction of the SSNMR pellet. Cell extracts were prepared by resuspending 100 mg of fresh wet cell pellet or the frozen-cell pellet in 200 μL fructose-supplemented liquid Schuster minimal media, then sonicating them (4 min, 5 s pulse on, 5 s pulse off; on ice) with a Fischer Scientific Model 550 Sonic Dismembrator. The cell extract samples appear homogeneous, translucent and yellow.
Resuspended cells and cell extracts were measured in a 50 mm stem coaxial insert with an external D₂O reference reservoir. All ³¹P solution NMR spectra were measured at room temperature (ca. 293 K) with a 5 s recycle delay, and 5200 scans were averaged (8 h per spectrum) on a Bruker AVANCE NEO 400 MHz spectrometer.