P‐31 Nuclear Magnetic Resonance Analysis of Brain: The Perchloric Acid Extract Spectrum

Abstract

Abstract: Perchloric acid (PCA) extracts were prepared from liquid‐N 2 ‐frozen guinea pig brains and their organophosphate profiles examined by P‐31 nuclear magnetic resonance (NMR) spectroscopy. Thirty‐two phosphorus‐containing brain metabolites were characterized and quantitated. A distinctive feature of brain tissue metabolism relative to that of other tissues probed by P‐31 NMR is its pronounced ribose 5‐phosphate content. Comparison of brain metabolite levels following control or sublethal cyanide treatment (4 mg/kg) revealed specific cyanide‐induced changes in brain metabolism. Brains from cyanidetreated animals were characterized by a reduced phosphocreatine content and elevated α‐glycerolphosphate and inorganic orthophosphate contents relative to control. P‐31 NMR spectra of brain PCA extracts at pH 7.2 were also obtained under conditions that approximate those used for in vivo and intact tissue in vitro P‐31 spectroscopic analyses. The spectra reveal nine separate resonance bands corresponding to: sugar phosphates, principally ribose 5‐phosphate (3.7δ); inorganic orthophosphate (2.2δ); glycerol 3‐phosphorylethanolamine (0.3δ); glycerol 3‐phosphorylcholine (−0.1δ); phosphocreatine (−3.2δ); adenosine tri‐(β‐ATP) and di‐(β‐ADP) phosphate ionized end‐groups (−6.2δ); α‐ATP, α‐ADP, and nicotinamide adenine dinucleotides esterified end‐groups (−11.1δ); uridine diphosphohexose, hexose esterified end‐groups (−13.0δ); and β‐ATP ionized middle group (−21.6δ). Knowledge of the phosphatic molecules that contribute resonances to the brain P‐31 NMR spectrum as well as understanding their magnetic resonance properties is essential for the interpretation of in vivo brain spectroscopic data as well as brain extract data, since these same compounds contribute to the intact brain P‐31 spectrum.