Measurements of macromolecule-bound and ultra-filtrable polyamines in rat liver homogenized without buffer

Ultra-filtrable and macromolecule-bound polyamines in rat liver homogenates , made without buffer, were determined, using Potter-Elvehjem homogenizer a nd commercially available, pressure-aided ultrafiltration device with a mem brane pore size that allows passage of particles of molecular weight no lar ger than 5000, About 90% of polyamines in the liver were shown to be equili brated with externally added N-15-labeled polyamines, based on the differen ce in the ratio of the natural to N-15-labeled polyamine in the liver homog enate and the ultrafiltrate, The entire amount of ultrafiltrate in the homo genized liver, required for calculation of the amounts of ultra-filtrable a nd macromolecule-bound polyamines, was estimated to be about 0.25 g in one gram of the homogenate, using a limited dilution curve of spermine in the u ltrafiltrate with phosphate buffered saline and distilled water. With this value, ultra-filtrable polyamines in normal rat liver homogenate were calcu lated as about 25%, 8%, and 2% of the total amount of putrescine, spermidin e, and spermine, respectively. The method was then used to measure ultra-fi ltrable and macromolecule-bound polyamines in regenerating rat liver homoge nates, to examine possible changes of polyamines during cell growth. The me thod was also applied to measure other ultra-filtrable compounds such as am ino acids and inorganic ions in rat liver homogenate.