Details

Autor(en) / Beteiligte

• Egan, B.Zane
• Rhear, R.W.
• Kelmers, A.D.

Titel

Solvent extraction of transfer ribonucleic acids

Ist Teil von

• Analytical biochemistry, 1970-11, Vol.38 (1), p.139-147

Ort / Verlag

United States: Elsevier Inc

Erscheinungsjahr

1970

Quelle

Elsevier Journal Backfiles on ScienceDirect (DFG Nationallizenzen)

Beschreibungen/Notizen

• Mixed tRNA's from E. coli are selectively extracted from an aqueous solution containing 0.05 M Tris-HCl, 0.01 M MgCl 2, and various concentrations of NaCl with an organic solution containing Adogen 464, a tetraalkylammonium chloride, in Freon 214. The tRNA's are completely extracted into the organic phase at NaCl concentrations below 0.1 M, and the extraction coefficient [ E α 0 = A 260 ( org) A 260 ( aq) ] decreases with increasing NaCl concentration. The extraction coefficient increases with increasing temperature in the range 10° to 25°C. Consequently, the tRNA's can be stripped from the organic phase with an aqueous solution of higher salt concentration, e.g., 0.5 M NaCl, or at lower temperature, e.g., 5°C. Other parameters influencing the extraction coefficient are extractant concentration in the organic phase and Mg 2+ concentration in the aqueous phase. With other conditions constant, the extraction coefficients for 0.1 M and 0.5 M amine solutions are 3.75 and 15.1, respectively. The usable extraction range occurs at higher NaCl concentrations in the absence of MgCl 2. The order of extractability of the specific tRNA's can be correlated with the order of elution from a reversed-phase chromatographic column. The tRNA's that elute first are also generally the least extractable, while those that elute last are the most extractable. A tRNA Phe product having 399 pmoles A 260 unit has been prepared by a simple batch-type extraction. The major active contaminants were tRNA Tyr ( 75 pmoles A 260 unit ) and tRNA Trp ( 56 pmoles A 260 unit ). It was possible to obtain a product which assayed approximately 800 pmoles A 260 unit for tRNA Phe by combining batch extraction with gel permeation chromatography.