Partitioning of Some Amino Acids and Low Molecular Mass Peptides in Aqueous Two-Phase Systems of Poly(ethylene glycol) and Dextran in the Presence of Small Amounts of K₂HPO₄/KH₂PO₄-Buffer at 293 K:  Experimental Results and Correlation

Abstract

Buffered aqueous two-phase systems are effective extraction systems for separating amphoteric hydrocarbons like, for example, polypeptides from aqueous phases. The design and basic engineering of such processes require the knowledge of the liquid-liquid equilibrium. Methods to calculate and predict the liquid-liquid phase equilibrium in those systems can only be developed when a large data base is available. The present contribution is aimed at that goal by presenting experimental results for the partitioning of small amounts (≈0.001 g/g solution) of model substancessamino acids glycine, L-glutamic acid, L-phenylalanine, and L-lysine as well as dipeptides and some higher, but still low, molecular peptides of those single amino acidssin aqueous two-phase systems of dextran and poly(ethylene glycol) in the presence of small amounts (about 0.05 mol/kg) of K 2 HPO 4 /KH 2 PO 4 -buffer at about 293 K. A group contribution model for the excess Gibbs energy is used to correlate/predict that liquid-liquid phase equilibrium.

Partitioning of low molecular combination peptides in aqueous two-phase systems of poly(ethylene glycol) and dextran in the presence of small amounts of K2HPO4/KH2PO4 buffer at 293 K: Experimental results and predictions

“…Lower phase Upper phase From a previous publication (Tintinger et al, 1997b), it is known that the amino acids glycine, L-glutamic acid, and L-lysine prefer the DEX-rich lower phase over the PEG-rich upper phase, whereas L-phenylalanine shows the inverse preference. From that publication it is also known that peptides consisting of either (only) glycine, glutamic acid, or lysine groups also prefer the DEX-rich over the PEG-rich phase while peptides of (only) phenylalanine groups enrich in the PEG-rich phase.…”

“…When the concentrations of PEG and DEX were not analysed they were taken from the literature (Großmann et al, 1995b). As in a previous publication (Tintinger et al, 1997b), the partitioning of the buffer was not determined exactly, as only the potassium ion concentration was measured in both coexisting phases. The experimental uncertainty of the potassium ion molality in a phase is about 6%.…”

“…The present publication is a supplement to previous work (Großmann et al, 1993(Großmann et al, , 1995a(Großmann et al, ,b, 1997Tintinger et al, 1997a,b) on the liquid-liquid phase equilibrium of aqueous two-phase systems and the partitioning of amino acids and peptides. The last publication of that series (Tintinger et al, 1997b) deals with the partitioning of small amounts (≈10 −3 g substance per gram solution) of amino acids glycine (gly), L-glutamic acid (glu), Lphenylalanine (phe), and L-lysine (lys) and some of their low molecular peptides (all containing only groups from a single amino acid) as e.g. dipeptides gly-gly, glu-glu, phephe, and lys-lys and tripeptides 3*gly, 3*glu, and 3*lys in aqueous two-phase systems of PEG 6000 (or PEG 35000) and DEX 500 at 293 K buffered to pH ≈ 7 by adding small amounts (≈ 0.05 mol/kg solution) of an equimolar buffer of K 2 HPO 4 and KH 2 PO 4 .…”

“…That model is an osmotic virial equation which combines the idea of splitting molecules into groups (introducing group contributions) with the idea of using surface fractions to encounter for the probability of intermolecular interactions (Großmann et al, 1995a,b;Großmann, 1994). That model has been used to correlate/predict the partitioning of amino acids and peptides (of single amino acids) (Tintinger et al, 1997b). From those correlations all model parameters were adopted for the predictions of the present work.…”

“…Table I gives information on the chemicals used in the present investigation as well as on some of their thermophysical properties needed for phase equilibrium calculations. Polymer samples (poly(ethylene glycol) PEG 6000 and PEG 35000 and dextran DEX 500) were the same as in previous work (Tintinger et al, 1997b). The number averaged molecular mass M n of the poly(ethylene glycols) is M n (PEG 6000) ‫ס‬ 6,230 and M n (PEG 35000) ‫ס‬ 34,100.…”