Phospholipase A2-Catalyzed Membrane Leakage Studied by Immobilized Liposome Chromatography with Online Fluorescent Detection

Effect of lipid composition on phospholipase A2‐catalyzed membrane leakage in immobilized liposomes: Sensitization for polychlorinated biphenyls detection with antibody affinity column tandem with fluorescent liposome column

“…1A), similar to our previous report ( Fig. 1 in [12]. The similarity of PLA 2 -catalyzed membrane leakage using fluorescent on-line and off-line analysis indicated that the immobilized liposomes packing in the open column could also simply and precisely determine the calcein release by PLA 2 hydrolytic action.…”

“…time, the enzyme has sufficient time to diffuse into the gel beads and efficiently hydrolyze the immobilized liposomes. This result is different from our previous report of the effect of flow rate on PLA 2 hydrolytic action by ILC-fluorescent on-line analysis [12], which is not time dependent in the chromatographic flow rate range of 0.1,1.0 mL/min. Moreover, it was found that the fluorescent signal can return to the baseline quickly when only 4 mL of elution buffer was applied to the 20 min incubated column compared with that on the 1 or 5 min incubated liposome column.…”

“…In our previous work [12], we found that the largest PLA 2 -induced calcein leakage from the liposome column could be detected on LUV-TSK G6000PW or -Sephacryl S1000 gel in the presence of 1 mM Ca 21 in the mobile aqueous phase. In this work, Sephacryl S1000 gel was used because the particle size of TSK G6000PW gel is too small to pack in the Bio-Spin open column.…”

“…3 Results and discussion 3.1 Open liposome column analysis of PLA 2 -catalyzed membrane leakage PLA 2 -catalyzed membrane leakage can be analyzed by ILC with on-line fluorescent detection [12]. In the open liposome column analysis, the calcein release increased with increasing the enzyme concentration (Fig.…”

“…An immobilized liposome column (ILC) has been developed for various chromatographic applications, such as immobilized liposomes containing a self-quenching fluorescent dye, calcein, for bioanalysis [11,12]. The advantage of the fluorescent liposome column is its excellent stability upon storage and chromatographic runs [11].…”