A multi stage extraction procedure which gives the possibility to analyze both water soluble and lipid components stemming from the same specimen has been developed in our laboratory. Metabolites from brain cancer biopsies have been compared to metabolites in normal human brain, developing mouse brain and primary mouse cell cultures of neurons and astrocytes from cerebral cortex and cerebellum. Extraction with perchloric acid (PCA) dissolves water.soluble components such as choline containing compounds, creatine, amino acids, carbohydrates and high energy phosphates. The water insoluble fraction left after the PCA treatment was extracted with chloroform/methanol, 2/1 (vol/vol). C-13 and H-1 NMR spectra showed characteristic lipid resonances identified as those from long chain saturated and unsaturated fatty acids and cholesterol. Only glioblastomas contained detectable amounts of cholesteryl ester suggesting this compound as marker for brain pathology. It was shown that cholesterol but not cholesteryl ester is present in cultures of neurons either alone or together with astrocytes. H-1 NMR spectra of PCA extracts from biopsies showed that the creatine/choline containing compounds (Cr/Cho) ratios decreased and the N-acetylaspartate/Cho ratios decreased in glioblastomas as compared to normal brain. Cell cultures retain the Cr/Cho ratio characteristic of the developmental stage of the tissue they were prepared from.

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