Simple and Efficient Profiling of Phospholipids in Phospholipase D-modified Soy Lecithin by HPLC with Charged Aerosol Detection

• Published in: Journal of the American Oil Chemists' Society 2013-07, Vol.90 (7), p.951-957
• Main Authors: Damnjanović, Jasmina, Nakano, Hideo, Iwasaki, Yugo
• Format: Article
• Language: English
• Subjects: Aerosols; Agriculture; Biomaterials; Biotechnology; Charged aerosol detector; Chemical reactions; Chemistry; Chemistry and Materials Science; Chloroform; detection limit; Enzymes; Food Science; high performance liquid chromatography; HPLC; Industrial Chemistry/Chemical Engineering; Lipids; Liquid chromatography; Original Paper; phospholipase D; Phospholipids; Silica; solvents; Soy lecithin; Soybeans; Streptomyces
• ISSN: 0003-021X; 1558-9331
• DOI: 10.1007/s11746-013-2236-x

Dietary phosphatidylinositol (PI) can be synthesized via phospholipase D (PLD)-catalyzed transphosphatidylation of phosphatidylcholine (PC), abundant in soy lecithin, with myo-inositol. However, a generated mixture of phospholipid (PL) classes poses a challenge for analysis. Our current work on Streptomyces PLD engineering requires a robust analytical method for profiling of PI and related PLs derived from the transphosphatidylation reactions. Therefore, we optimized an HPLC-based method with charged aerosol detector (CAD) for PL quantification. PLs were separated on a normal phase silica column by a gradient elution system using two solvents containing chloroform/methanol/1 M formic acid–triethylamine buffer in different ratios. Retention times of the PL standards and LC–MS under identical conditions were used to identity PL classes. PL standards gave linear response in 100- and 10-fold (lyso-PI) concentration range. The method provided a simple, sensitive, repeatable, and precise analysis of PI, PC, phosphatidylethanolamine, phosphatidic acid, and lyso forms of PC and PI. Compared to the similar existing method, introduction of CAD provided a three- to fivefold decrease at the lower end and a two- to fivefold increase at the upper end of the dynamic range. High precision, high sensitivity, and low limits of detection and quantification further underline the benefits of CAD in PL analysis.