Polypropylene microcentrifuge tubes

Briefly, bacteria broth was collected after cultivation overnight in the absence or presence of individual beta-lactams, rinsed with 1x PBS, spun and dried with a speed-vac. 150 μL of water was then added to the pelleted and dried bacteria. The resulting suspensions were sonicated in an ice bath for 30 min to dislodge the dried pellets and homogenize the suspension. A chilled solution of chloroform and methanol (1:2 v/v, 600 μL) was added to each tube, followed by 5 min of vortex and the addition of 150 μL of chilled chloroform and 150 μL of chilled water. The samples were then rigorously vortexed for 1 min and centrifuged for 10 min at 4°C and 2,000 × g to separate the organic and aqueous layers. The organic layers were collected to clean 1.5 ml polypropylene microcentrifuge tubes (Fisher Scientific, Waltham, MA, United States) and dried in a vacuum concentrator. The dried lipid extracts were reconstituted with 500 µL of 2:1 acetonitrile/methanol and transferred to glass vials for storage prior to LC-MS analysis.