Cation-exchange gradient

Lysozyme and ribonuclease were separated by cation-exchange gradient elution (both strong and weak ion dtange columns) as a function of gra dient time and flow rate (2S). Values of Z were obtained from isocratic data (Fig. 8) and used to predict Ox values for these gradient runs. There data are summarized in Table XI. Overall agreement is acceptable, with 0.98 and SD 0.17 for 14 ta points. 

The applicability of induced pH gradients for separation of metal ions within anion- and cation-exchange columns was verified. 

Fig. 3. Cation-exchange chromatography of protein standards. Column poly(aspartic acid) Vydac (10 pm), 20 x 0.46 cm. Sample 25 pi containing 12.5 pg of ovalbumin and 25 pg each of the other proteins in the weak buffer. Flow rate 1 ml/min. Weak buffer 0.05 mol/1 potassium phosphate, pH 6.0. Strong buffer same +0.6 mol/1 sodium chloride Elution 80-min linear gradient, 0-100% strong buffer. Peaks a = ovalbumin, b = bacitracin, c = myoglobin, d = chymotrypsinogen A, e = cytochrom C (reduced), / = ribonuclease A, g = cytochrome C (oxidised), h = lysozyme. The cytochrome C peaks were identified by oxidation with potassium ferricyanide and reduction with sodium dithionite [47]...

In Fig. 11, separations of chymotrypsinogen A, cytochrome c and lysozyme on strong cation exchangers carrying SO3 groups by gradient elution are shown. 

Fig. 11a. Fractionation of (/) chymotrypsinogen A, (2) cytochrome C and (3) lysozyme on strong cation exchangers, a) Support Fractogel TSK 650(s)SP (conventional type) sample, 1 mg each flow rate, 1 ml/min column size, 150 x 10 mm T.D. Solvent A = 0.02 mol/1 phosphate, pH 6.0 solvent B = A + 1 mol/1 NaCl gradient, 0-10 min, 0% B 10-70 min, 0-100% B. b) Support Fractogel EMD 650(s)SO( — (tentacle type) conditions as in (a) [78]...

The dialysed sample was fractionated by cation exchange chromatography. A 40-50 ml sample was applied to a CM-Sepharose CL-6B column (1.5 x 15 cm). Unbound proteins were removed with 50 mM MES pH 6.8, 1 mM DTT, and the bound proteins were eluted with an increasing NaCl gradient from 0 - 0.4 M NaCl in a total volume of 500 ml. The flow was 25 ml/h and fractions of 8.33 ml were collected. The protein profile was measured at 280 nm. 

Gerstner, J. A. and Cramer, S. M., Cation-exchange displacement chromatography of proteins with protamine displacers effect of salt-induced gradients, Biotechnol. Prog., 8, 540, 1992. 

After sample loading, the cation-exchange RAM-column was placed in-line with the analytical cation-exchange column and analytes were eluted with a salt gradient. A total of 24 fractions of 4 min duration (2 mL of eluent) were transferred to the... 

Essader, A.S., Cargile, B.J., Bundy, J.L., Stephenson, J.L., Jr. (2005). A comparison of immobilized pH gradient isoelectric focusing and strong-cation-exchange chromatography as a first dimension in shotgun proteomics. Proteomics 5, 24—34. 

Fig. 20. Test of stability of weak cation exchange monolithic column (ISCO). Conditions column, 50 X4.6 mm i.d., mobile phase gradient of sodium chloride in 0.01 mol/1 sodium phosphate buffer (pH 7.6) from 0.1 to 0.5 mol/1 in 4.5 min and to 1 mol/1 in 6.5 min, overall gradient time 11 min, flow rate 10 ml/min. Peaks Ribonuclease (1), cytochrome c (2), lysozyme (3). The two separations shown in this figure were achieved 503 runs apart...

Figure 4.8 Cation-exchange liquid chromatography of basic proteins. Column, Asahipak ES502C eluent, 20 min linear gradient of sodium chloride from 0 to 500 mM in 50 mM sodium phosphate buffer pH 7.0 flow rate, 1 ml min-1 temperature, 30 °C detection, UV 280 nm. Peaks 1, myoglobin from horse skeletal muscle (Mr 17 500, pi 6.8-7.3) 2, ribonuclease from bovine pancreas (Mr 13 700, pi 9.5-9.6) 3, a-chymotrypsinogen A from bovine pancreas (Mr 257 000, pi 9.5) and 4, lysozyme from egg white (Mr 14 300, pi 11.0-11.4). (Reproduced by permission from Asahikasei data)...

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