Characterization of nonexchangeable radioactivity in L1210 cells incubated with ( sup 14 C)thiotepa: Labeling of phosphatidylethanolamine

Journal Article · Sun Jul 01 04:00:00 EDT 1990 · Cancer Research; (USA)

OSTI ID:6698760

Egorin, M J; Snyder, S W ^[1]

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1. Univ. of Maryland Cancer Center, Baltimore (USA)

N,N',N''-Triethylenethiophosphoramide ((14C)thiotepa) accumulation by L1210 cells is a biphasic process. A very rapid initial phase is followed by a much slower second phase that reflects accumulation of radioactivity in a form that is not lost or exchanged when cells are resuspended and incubated in drug-free medium for up to 8 h. In this study we attempted to characterize this nonexchangeable radioactivity. Nuclei (10(7)) isolated from L1210 cells and incubated with (14C)thiotepa did not accumulate 14C during incubations of up to 5 h. Similarly, nuclei isolated from 10(7) L1210 cells that had been shown to accumulate nonexchangeable 14C after incubation with (14C)thiotepa did not show an increase in nuclear-associated 14C. Eighty to 85% of nonexchangeable 14C in L1210 cells incubated with (14C)thiotepa was soluble in ethanol or chloroform:methanol (2:1, v/v), and although most of this cell-associated nonexchangeable 14C was precipitated by trichloroacetic acid, subsequent treatment of that precipitate with methanol solubilized most of the 14C so that only 15 to 20% remained with the final precipitate. When chloroform:methanol-soluble nonexchangeable 14C was analyzed with thin-layer chromatography systems suitable for thiotepa or simple lipids, all radioactivity remained at the origin. In contrast, when analyzed with one- and two-dimensional thin-layer chromatographic systems suitable for complex lipids, all chloroform:methanol-soluble radioactivity was associated with a single lipid spot. This lipid cochromatographed with phosphatidylethanolamine, reacted with ninhydrin but not with 4-(p-nitrobenzyl)pyridine or the Dragendorff choline reagent, and was digested by phospholipases C and D, all of which lead to its identification as phosphatidylethanolamine.

• OSTI ID: 6698760
• Journal Information: Cancer Research; (USA), Journal Name: Cancer Research; (USA) Vol. 50:13; ISSN 0008-5472; ISSN CNREA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
550901 -- Pathology-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ANIMAL CELLS
BIOLOGICAL ACCUMULATION
CARBON 14 COMPOUNDS
CELL CONSTITUENTS
CELL NUCLEI
CHEMICAL COMPOSITION
CHROMATOGRAPHY
DISEASES
ESTERS
HEMIC DISEASES
IMMUNE SYSTEM DISEASES
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
LABELLING
LEUKEMIA
LIPIDS
NEOPLASMS
ORGANIC COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
PHOSPHOLIPIDS
RADIOACTIVITY
SEPARATION PROCESSES
THIN-LAYER CHROMATOGRAPHY
TRACER TECHNIQUES
TUMOR CELLS