Separation of representative lipid compounds of biological membranes and lipid derivatives from peroxidized polyunsaturated fatty acids by reversed phase high-performance liquid chromatography.
A complex mixture of different lipid compounds, including phosphatidylcholine, phosphatidylserine, all trans-retinol, 15(S)-hydroperoxyeicosatetraenoic acid, D-alpha-tocopherol, saturated and unsaturated fatty acids can be separated by reversed phase HPLC by using a C-18, 120 mm x 4 mm, 3 microns particle size column and a step gradient from acetonitrile/water (1:1; v:v) to 100% acetonitrile at a flow rate of 0.8 ml/min. By applying this elution condition, separation of various groups of lipid hydroperoxides and lipid derivatives, each one originating from a different in vitro peroxidized polyunsaturated fatty acid, can be obtained. Simultaneous detection is carried out by a diode array detector at a wavelength accumulation range set up between 195 and 400 nm. The possibility of simultaneously having such a large number of measurements renders this chromatographic method particularly suitable in studies concerning lipid peroxidation where, in addition to the detection of free radical-induced lipid hydroperoxides, data on some key antioxidant molecules, i.e. vitamin A and E, as well as that of structural compounds of biological membranes, i.e. phosphatidylcholine and phosphatidylserine, can be achieved.
Di Pierro
D
D
Department of Experimental Medicine and Biochemical Sciences, University of Rome Tor Vergata, Italy.
Tavazzi
B
B
Lazzarino
G
G
Galvano
M
M
Bartolini
M
M
Giardina
B
B
D000097:Acetonitriles; D016718:Arachidonic Acid / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005231:Fatty Acids, Unsaturated / Q000302:isolation & purification*; D015289:Leukotrienes / Q000302:isolation & purification; D019787:Linoleic Acid; D008041:Linoleic Acids / Q000302:isolation & purification; D015227:Lipid Peroxidation; D008054:Lipid Peroxides / Q000302:isolation & purification*; D008055:Lipids / Q000302:isolation & purification*; D008563:Membrane Lipids / Q000302:isolation & purification*; D019301:Oleic Acid / Q000302:isolation & purification; D019308:Palmitic Acid / Q000302:isolation & purification; D010713:Phosphatidylcholines / Q000302:isolation & purification; D010718:Phosphatidylserines / Q000302:isolation & purification; D013229:Stearic Acids / Q000302:isolation & purification; D014801:Vitamin A / Q000302:isolation & purification; D014810:Vitamin E / Q000302:isolation & purification; D017965:gamma-Linolenic Acid / Q000302:isolation & purification
D000097:Acetonitriles; D005231:Fatty Acids, Unsaturated; D015289:Leukotrienes; D008041:Linoleic Acids; D008054:Lipid Peroxides; D008055:Lipids; D008563:Membrane Lipids; D010713:Phosphatidylcholines; D010718:Phosphatidylserines; D013229:Stearic Acids; D014801:Vitamin A; D014810:Vitamin E; D016718:Arachidonic Acid; D019301:Oleic Acid; D019308:Palmitic Acid; C031183:stearic acid; C025086:15-hydroperoxy-5,8,11,13-eicosatetraenoic acid; D017965:gamma-Linolenic Acid; D019787:Linoleic Acid; C032159:acetonitrile