PURPOSE: The weak aqueous interaction between the protein lactate dehydrogenase (LDH) and the nonionic surfactant Tween 80 has been investigated, because weak protein-amphiphile interactions are of significant importance in pharmaceutical formulations, but are experimentally hard to determine. The system LDH/sodium dodecyl sulphate (SDS) was used as reference because SDS, by its strong protein binding, denatures LDH completely. METHODS: Fluorescence spectroscopy with pyrene and 1,3-bis(lphenyl)propane (P3P) as probes, intrinsic protein fluorescence and NMR spectroscopy have been used. RESULTS: The fluorescence probe pyrene monitors a weak Tween-LDH interaction, detectable below the critical micelle concentration of ordinary Tween micelles. The microviscosity probe P3P shows a surfactant-induced denaturation in the case of LDH/SDS but not in the case of LDH/Tween 80. Intrinsic LDH fluorescence verifies this behavior. Pulsed-gradient spin-echo NMR was also used to verify the weak LDH-Tween 80 interaction. CONCLUSIONS. A weak interaction between LDH and Tween 80 occurs at hydrophobic zones of the protein, but it is not strong enough to denature LDH. The experimental outline used here provides a useful approach for mapping the very weak protein-amphiphile interactions often present in pharmaceutical formulations.
PURPOSE: The weak aqueous interaction between the protein lactate dehydrogenase (LDH) and the nonionic surfactant Tween 80 has been investigated, because weak protein-amphiphile interactions are of significant importance in pharmaceutical formulations, but are experimentally hard to determine. The system LDH/sodium dodecyl sulphate (SDS) was used as reference because SDS, by its strong protein binding, denatures LDH completely. METHODS: Fluorescence spectroscopy with pyrene and 1,3-bis(lphenyl)propane (P3P) as probes, intrinsic protein fluorescence and NMR spectroscopy have been used. RESULTS: The fluorescence probe pyrene monitors a weak Tween-LDH interaction, detectable below the critical micelle concentration of ordinary Tween micelles. The microviscosity probe P3P shows a surfactant-induced denaturation in the case of LDH/SDS but not in the case of LDH/Tween 80. Intrinsic LDH fluorescence verifies this behavior. Pulsed-gradient spin-echo NMR was also used to verify the weak LDH-Tween 80 interaction. CONCLUSIONS. A weak interaction between LDH and Tween 80 occurs at hydrophobic zones of the protein, but it is not strong enough to denature LDH. The experimental outline used here provides a useful approach for mapping the very weak protein-amphiphile interactions often present in pharmaceutical formulations.
Authors: M J Adams; G C Ford; R Koekoek; P J Lentz; A McPherson; M G Rossmann; I E Smiley; R W Schevitz; A J Wonacott Journal: Nature Date: 1970-09-12 Impact factor: 49.962
Authors: L Kreilgaard; L S Jones; T W Randolph; S Frokjaer; J M Flink; M C Manning; J F Carpenter Journal: J Pharm Sci Date: 1998-12 Impact factor: 3.534