Optimal Conditions for the Direct RP-HPLC Determination of Underivatized Amino Acids with Online Multiple Detection

Area of Science:

• Analytical Chemistry
• Biochemistry
• Chromatography

Background:

• Accurate detection of amino acids and related compounds is crucial in various scientific fields.
• Existing methods may require derivatization or lack comprehensive detection capabilities.
• There is a need for efficient, simultaneous analysis of multiple underivatized amino acids and related metabolites.

Purpose of the Study:

• To develop and validate a sensitive analytical method for the simultaneous detection of 15 underivatized amino acids, 2 dipeptides, and creatinine.
• To utilize a combination of dual-UV, fluorimetric, and multielectrode electrochemical detectors for enhanced detection capabilities.
• To establish an efficient separation protocol using gradient elution chromatography.

Main Methods:

• Employed a dual-UV detector, a fluorimetric detector, and a multielectrode electrochemical detector with a glassy carbon electrode and carbon microfibers.
• Separated a mixture of 18 underivatized solutes, including amino acids, dipeptides, and creatinine.
• Utilized a multilinear acetonitrile gradient elution program optimized by an algorithm for reversed-phase separation.

Main Results:

• Successfully detected 15 underivatized amino acids, 2 dipeptides (L-cystathionine and L-carnosine), and creatinine.
• Achieved efficient separation of the 18 solutes within a 27-minute timeframe.
• Demonstrated the combined utility of multiple detection techniques for comprehensive analysis.

Conclusions:

• The proposed method enables the simultaneous and sensitive detection of a wide range of underivatized amino acids and related compounds.
• The multielectrode electrochemical detector, particularly with microfibers, offers enhanced detection capabilities.
• This approach provides an efficient and effective analytical solution for complex biological mixtures.