31P NMR studies of cultured human tumor cells. Influence of ph on phospholipid metabolite levels and the detection of cytidine 5′‐diphosphate choline

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Abstract
202 MHz 31P NMR (11.7 T) was used to study the effects of culture medium pH on the levels of phosphate metabolites in three human tumor cell lines (XP29MAmal, a malignant xeroderma pigmentosum; CX-1, a colon carcinoma; KB, a squamous cell carcinoma of the oral cavity). Cells were cultured in Roux flasks in HAM's F-12 medium, and the pH was varied with the final medium change. After harvesting, 1–5 × 108 cells were suspended in Ringer/HEPES buffer at pH 7.4 and 4°C for 31P NMR studies. Cell adhesion and growth rate decreased with decreasing pH, but, down to ca pH 6.1, trypan blue exclusion and the observed levels of nucleoside di- and triphosphates (range: 22–37% of total phosphates detected), phosphocreatine (PCr, 2–5%) and Pi (5–11%) did not vary significantly with pH. For XP29MAmal cells in exponential growth phosphocholine levels decreased from 18–28% at pH 7.0 to ca 5% at pH 6.0, while phosphoethanolamine levels increased from 2–7% to 15%. Glycerophosphocholine (GPC) levels increased from ca 7% at pH 7.2 to 13% at pH 6.3. At pHP: Pα = −8.13 ppm, Pβ = 8.93 ppm, for PCr = 0 ppm). However, confluent cells did not accumulate CDP-choline when the pH was lowered. The cell lines CX-1 and KB also showed the pH effects described herein.