Fractionation of soybean phospholipids by preparative high-performance liquid chromatography with sorbents of various particle size.
Normal-phase high-performance liquid chromatography was used on a preparative scale to seperate phosphatidylethanolamine (PE), phosphatidylinositol (PI) and phosphatidylcholine (PC) phospholipids from soybean. Separation was achieved using mixtures of three solvents, hexane, methanol and isopropanol. The optimized mobile phase compositions were experimentally determined while operating in a linear gradient mode using 15, 5-20, 25-40, and 40-63 microm preparative particles as well as 4 microm analytical particles. A gradient mobile phase was established on a commmercially available analytical Nova-Pak column such that hexane linearly decreased from 85 to 0 as isopropanol and methanol linearly increased in two gradient steps from 10 to 30 and 5 to 70 respectively. The total run time was 25 min at a flow-rate of 1.5 ml/min. A slight change in mobile phase composition was required to increase the resolution of phospholipids. The 15 microm particle size gave the best separation of the preparative particle sizes examined based on their resolutions between PE and PI and PI and PC. Finally, the retention factors of PE and PC were correlated in terms of mobile phase composition.
Identification of chiro-inositol-containing phospholipids and changes in their metabolism upon salt stress in soybean seedlings.
Abstract We report here the presence of chiro-inositol-containing phospholipids in addition to the common myo-inositol-containing phospholipids in soybean seedlings. Using sequential TLC and HPTLC, eleven inositol phospholipids were isolated from soybean seedlings and analyzed after 6 N HCl hydrolysis for their inositol and fatty acid compositions by anion exchange Dionex HPLC and GC, respectively. Upon analysis, we have identified three species of chiro-inositol-containing phospholipids and further observed that the chiro-inositol-containing phospholipids and their fatty acid composition changed in response to 1 and 24 h 150 mM NaCl treatment compared to the control (no NaCl treatment). In addition, we found the different tissue distribution and metabolic changes of the chiro-inositol-containing phospholipids in apical, elongating and mature parts of hypocotyls and roots of soybean seedlings upon salt stress.
Sterol metabolism. XLI. Cholesterol A-ring autoxidations
Abstract Chromatographic and spectral evidence is adduced for the presence of cholest-5-en-3-one, cholest-4-en-3-one, and cholest-4-ene-3,6-dione in samples of cholesterol aged naturally in air or subjected to irradiation in air by 60 Co gamma radiation. These findings establish an additional mode of air oxidation of cholesterol to A-ring 3-ketones. Moreover, the oxidation by air of cholest-5-en-3-one induced by 60 Co gamma radiation yielded cholest-4-en-3-one, cholest-4-ene-3,6-dione, and the epimeric 3-oxocholest-4-ene-6-hydroperoxides. Cholest-4-en-3-one was not altered by irradiation in air, nor was cholesterol isomerized to cholest-4-en-3β-ol upon irradiation. From these observations it is deduced that the radiation-induced A-ring dehydrogenation of cholesterol yields initially cholest-5-en-3-one which upon isomerization yields cholest-4-en-3-one not further oxidized and which by a second oxidation yields the epimeric 3-oxocholest-4-ene-6-hydroperoxides which decompose to cholest-4-ene-3,6-dione.