ABSTRACIThe addition of an elicitor (glucan) to Phaseolus vulgaris cell suspension cultures increased the formation of the phytoalexin phaseollin. Intracellular pH and phosphate concentrations were studied with 31P nuclear magnetic resonance spectroscopy on elicitor-treated cells which were aerated during the nuclear magnetic resonance measurement. The pH of the vacuole and to a lesser extent the pH of the cytoplasm were affected at 10 minutes after elicitor addition; a decrease in pH from 53 to 4.8 was noted in the vacuole and from 7A6 to 7.28 in the cytoplasm. The ratio between the amount of Pi in the vacuole to that in the cytoplasm also changed within 10 minutes after elicitor addition. The signal for ATP (8-ATP) was low after elicitor addition and was high again 23 hours after elicitation. tions and pH observed in plant cells after elicitor addition (24). In that study the plant cells were not aerated during the NMR measurements. It was shown that the amounts of inorganic phosphate in the vacuole increased at the expense of the phosphate in the cytoplasm of P. hortense cells after the addition of fungal cell wall fragments. It was briefly mentioned that the vacuolar pH decreased from 5.5 to 5.1 within 1 h after elicitor addition.The present study was carried out to establish more definitely the changes in cytoplasmic and vacuolar pH after the addition of an elicitor to P. vulgaris cell suspension cultures. These cells were studied under aerated conditions in the NMR tube. MATERIALS AND METHODSGrowth Conditions. Cells of Phaseolus vulgaris strain BRTIL-WAX (commercial) were maintained in the dark on SH medium (23) containing: 2,4-D, 0.5 mg/L; p-chlorophenoxyacetic acid, 2 mg/L; kinetin, 0.1 mg/L; nitrilotriacetic acid, 40 mg/L; and enzymatic casein hydrolyzate (Sigma, Cat. No. C-0626, St. Louis, MO), 2 g/L. An inoculum (10% by volume) of homogeneous suspension was transferred every 5 d to fresh medium. For 31P NMR experiments cells were transferred to SH medium without auxins (2,4-D and pCPA) and without the following medium components: Na7EDTA, nitrilotriacetic acid, FeSO4. 7H20, CoC12* 6H20, Na2MoO4 * 2H20, CuS04. 6H20, ZnSO4* 7H20, H3BO3, and MnSO444H20. Sodium chloride (20 mg/L) was added to this medium. All experiments were performed in 250 ml cotton plugged Erlenmeyer flasks with 100 ml medium. Growth was in a rotary shaker (G-25