Partitioning and antioxidant action of the water-soluble antioxidant, Trolox, between the aqueous and lipid phases of phosphatidylcholine membranes: 14C tracer and product studies.
The water-soluble antioxidant, 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic (Trolox), (4-14C)-labelled, was used to trace its location in the aqueous and lipid phases of liposomes. Trolox was found to partition 20 to 25% into the lipid phase of multilamellar (MLV) and 38-46% into the lipid phase of unilamellar (ULV) egg lecithin liposomes. Trolox and its oxidation products partition readily (40%) into the lipid phase of dilinoleoylphosphatidylcholine (DLPC) MLV liposomes during inhibited peroxidation, thermally initiated by azo-bis(2,4-dimethylvaleronitrile) (ADVN). The time-course of the consumption of Trolox during peroxidation of DLPC liposomes, initiated by ADVN, was followed by separation and analyses of [4-14C]Trolox and its oxidation products. Such studies showed that the consumption of Trolox followed the profile of the inhibition of oxygen uptake. This indicates that Trolox can be used in quantitative studies of membrane peroxidation; for example, to measure the rate of chain initiation (Ri). The product distribution of hydroperoxides, the 9- and 13-cis,trans (c,t) and trans,trans (t,t) isomers, formed during inhibited peroxidation of linoleate, in DLPC and methyl linoleate in dimyristoyl PC (DMPC) liposomes was determined by HPLC of the derived hydroxy methyl esters. The c,t/t,t (kinetic/thermodynamic) ratios were related to the antioxidant activity of the inhibitors. Both Trolox and alpha-tocopherol (vitamin E) gave relatively high initial c,t/t,t ratios (6.6 and 7.1) during inhibited peroxidation of DLPC, initiated by water-soluble azo-bis(2-amidinopropane.HCl) (ABAP). High initial c,t/t,t ratios (6.2) were also observed for alpha-tocopherol-inhibited peroxidation of DLPC liposomes, initiated by lipid-soluble ADVN. On the other hand, the combination of Trolox with ADVN-initiated peroxidation of DLPC or of methyl linoleate in DMPC gave relatively low initial c,t/t,t ratios of 3.5 and 1.3. These results are interpreted in terms of the relative hydrogen atom donating ability of the antioxidants and the homogeneity of the system used. The 9/13 ratios of hydroperoxides were constant (0.9 to 1.0) in all experiments and did not give evidence for preferential trapping by Trolox of peroxyls at the 9-position.
Barclay
L R
LR
Chemistry Department, Mount Allison University, Sackville, N.B., Canada.
Artz
J D
JD
Mowat
J J
JJ
D000975:Antioxidants / Q000737:chemistry*; D002250:Carbon Radioisotopes; D002839:Chromans / Q000737:chemistry*; D015227:Lipid Peroxidation; D008055:Lipids; D008081:Liposomes / Q000737:chemistry*; D010713:Phosphatidylcholines / Q000737:chemistry*; D014867:Water
D000975:Antioxidants; D002250:Carbon Radioisotopes; D002839:Chromans; D008055:Lipids; D008081:Liposomes; D010713:Phosphatidylcholines; D014867:Water; C010643:6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid