The Application Notebook
Volume 35, Issue 9
Amino acids have significant biological importance as they are the building blocks of life, they are also used in nutritional supplements, fertilizers, and food technology. Industrial amino acid use includes the production of drugs, biodegradable plastics, and even chiral catalysts. With the wide variety of uses, there are many ways to classify amino acids including polarity, pH, size, hydrophobicity, side chain group type, and so on. However the ideal, quantitative analysis of amino acids combines speed and sensitivity with reliability of the analytical technique.
Many columns cannot offer a single method to analyze the 20 building block amino acids. Shodex introduces the new HILICpak VC-50 2D column, which can offer speed, sensitivity, and reproducibility. This column is packed with polyvinyl alcohol material with carboxyl functional groups. When used according to the protocol described below, the column enables the user to analyze all 20 amino acids using a single method. A single run can be completed in 25 min with good resolution. In addition to analyzing the essential amino acids, the VC-50 2D has been shown to separate amino sugars.
Each amino acid was dissolved in water and diluted to 0.1 µg/mL. The analysis of all 20 amino acids was successfully accomplished with the Shodex HILICpak VC-50 2D (2.0 mm ID × 150 mm), a polymer based column for HILIC analysis. From each sample, 5 µL was injected into the system. The column temperature was kept at 30 °C and a flow rate of 0.2 mL/min. Eluent conditions were H2O/100 mM HCOOH aq./CH3CN = 30/20/50 to 0/90/10 (0 to 5 min), 0/90/10 (5 to 30 min) a lower pressure linear gradient. The HPLC system was coupled with an ESI-MS SIM detector.
The Shodex team has successfully developed a method for detecting 20 amino acids using a single method on the HILICpak VC-50 2D under positive MS mode. From the data shown above, isoleucine and leucine have some overlap. Through some modifications to the method, this analysis can result with a better separation. The Shodex HILICpak VC-50 2D is an ideal column for amino acid analysis.
Shodex™/Showa Denko America, Inc.
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