Quantification of Prostate Biopsy Metabolites

As previously described [19 (link)], ultrasound-guided prostate biopsies were acquired from 30 patients [15 from benign regions of the peripheral zone, 11 from regions of low-grade (Gleason score ≤ 3 + 4) cancer, and 4 from regions of high-grade (Gleason score ≥ 4 + 3) cancer] after approval by the UCSF Institutional Review Board (IRB) and informed consent using an 18- gauge needle (15-mm X 1-mm cores). Biopsies were placed in individual cryovials and snap-frozen on dry ice (≤15 s) immediately after the procedure. The tissues were processed for HR–MAS NMR, as described by Tessem et al. [40 (link)]. Briefly, each biopsy was weighed prior to loading into an HR–MAS rotor with a known mass of 3.0 μL of deuterium oxide containing 0.75% weight/volume sodium-3- trimethylsilylpropionate-2,2,3,3-d4 (D₂O + TSP). Quantitative ¹H HR–MAS NMR was performed at 11.7 T Varian INOVA NMR with a 4-mm gHX nanoprobe (Agilent Technologies, Santa Clara, CA, USA ) at 1 °C, and 2250 Hz spin rate. Fully relaxed pulse-acquire spectra were acquired with a 2-s presaturation delay, 2-s acquisition time, 40,000 points, 20,000-Hz spectral width, and 128 transients. The data were processed and quantified using the previously published HR–QUEST technique using ERETIC, an electronic reference to access ex vivo concentrations [60 (link)], yielding absolute concentrations of lactate [40 (link)].