Investigations of vacuoles isolated from tobacco: I. Quantitation of nicotine.

Nicotine was shown to be associated with mature vacuoles isolated from protoplasts of Nicotiana rustica. The vacuolar preparations also contained high levels of acid phosphatase, ATPase, and approximately 30% of the soluble protoplastic protein. The contamination of the vacuolar isolate by chlorophyll, succinate dehydrogenase, and NADPH cytochrome c reductase (markers for chloroplasts, mitochondria, and endoplasmic reticulum) was low. The enzymic activity associated with the vacuoles was not due to the exogenously supplied digestive enzymes used in the preparation of the protoplast. The relatively easy isolation of tobacco vacuoles makes this an excellent system for biochemical investigations of the vacuole.