FIGURE 2.

Detection of prenyl-pABA, DMQ₆, and Q₆ in lipid extracts of wild-type yeast cultured in the absence or presence of ¹³C₆-pABA. Wild-type yeast (W303-1A) were pre-cultured in Dogal media (minus pABA and folate) as described under “Experimental Procedures.” Cells were harvested during mid-log from large cultures (300–600 ml of media; A_{600 nm} = 0.6), resuspended in 2 ml of fresh media or fresh media containing 2–10 μg/ml [¹³C₆]pABA (final concentration), incubated at 30 °C and harvested after 16 h. Lipid extracts were prepared and quinones and prenylated intermediates were subjected to RP-HPLC-ESI-MS/MS as described. A–F show product ion spectra: A, prenyl-pABA [M+H]⁺ precursor ion (C₃₇H₅₅NO₂⁺; exact mass, 546.4) and the prenyl-pABA tropylium ion [M]⁺ (C₈H₈N₁O₂⁺; exact mass, 150.0); B, prenyl-[¹³C₆]pABA [M+H]⁺ precursor ion (¹³C₆¹²C₃₁H₅₅NO₂⁺; exact mass, 552.4) and the ¹³C₆-prenyl-pABA tropylium ion [M]⁺ (¹³C₆¹²C₂H₈NO₂⁺; exact mass, 156.0); C, DMQ₆ [M+H]⁺ precursor ion (C₃₈H₅₆O₃⁺; exact mass, 561.4) and the DMQ₆ tropylium ion [M]⁺ (C₉H₁₁O₃⁺; exact mass, 167.1); D, [¹³C₆]DMQ₆ [M+H]⁺ precursor ion (¹³C₆¹²C₃₂H₅₆O₃⁺; exact mass, 567.4) and the [¹³C₆]DMQ₆ tropylium ion [M]⁺ (¹³C₆¹²C₃H₁₁O₃⁺; exact mass, 173.1); E, Q₆ [M+H]⁺ precursor ion (C₃₉H₅₈O₄⁺; exact mass, 591.4) and the Q₆ tropylium ion [M]⁺ (C₁₀H₁₃O₄⁺; exact mass, 197.1); F, [¹³C₆]Q₆ [M+H]⁺ precursor ion (¹³C₆¹²C₃₃H₅₈O₄⁺; exact mass, 597.4) and the [¹³C₆]Q₆ tropylium ion [M]⁺ (¹³C₆¹²C₄H₁₃O₄⁺; exact mass, 203.1). Q₆ (Sigma) and the lipid extract prepared from NM101 (coq7-1) yeast strains were used to establish the retention times of Q₆ and DMQ₆, respectively (29). A, C, and E show the structures of the compounds detected and indicate the identity of the predicted tropylium-like ion of the base peak (tallest peak) formed under collision associated dissociative conditions.