Determination of Branched-Chain Amino Acids in Food Supplements and Human Plasma by a CE-MS/MS Method with Enhanced Resolution

Abstract

In the presented study, a capillary electrophoresis-mass spectrometry method combining high separation efficiency and sensitive detection has been developed and validated, for the first time, to quantify branched chain amino acids (valine, isoleucine, leucine) in commercial food and sport supplement samples and human plasma samples. The separations were performed in a bare fused silica capillary. The background electrolyte was composed of 500 mM formic acid with pH 2.0. The plasma sample pretreatment was realized by simple protein precipitation with acetonitrile. Injection of a short zone of highly basic electrolyte before the sample injection and application of the negative pressure on the separation were accompanied by enhanced resolution of the isobaric amino acids—isoleucine and leucine. The developed method was characterized by favorable validation parameters, such as linearity (r2 > 0.99), accuracy and precision, the limit of detection, lower limit of quantification, or robustness. These parameters were more than sufficient for the quantification of branched chain amino acids in various samples. The determined concentrations of branched chain amino acids in food and sports supplements were in very good agreement with the content declared by the manufacturer. The investigated concentrations of branched chain amino acids were in the range 294.68–359.24 µM for valine, 91.76–95.67 µM for isoleucine, and 196.78–251.24 µM for leucine. These concentrations fall within the physiological limits. The developed CE-MS/MS method represents a suitable alternative to traditional approaches used in branched chain amino acid quality control and bioanalysis.

The Role of Branched-Chain Amino Acids and Branched-Chain α-Keto Acid Dehydrogenase Kinase in Metabolic Disorders

“…A liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI ± -MS/MS) method makes it possible to investigate BCAAs accompanied by BCKAs in human serum simultaneously (10). The CE-MS method (9,(16)(17)(18) is a valuable alternative for detecting BCAA concentrations. This method is widely used in the quantification of BCAAs in fluids and food, also, it has a wide application in pharmaceutical quality control (16).…”

“…The CE-MS method (9,(16)(17)(18) is a valuable alternative for detecting BCAA concentrations. This method is widely used in the quantification of BCAAs in fluids and food, also, it has a wide application in pharmaceutical quality control (16).…”

Overview of chromatographic and electrophoretic methods for the determination of branched‐chain amino acids

“…Peak height was enhanced sixfold under pH-mediated stacking conditions [196]. Piestansky et al [197] performed CE-ESI-MS using an in-capillary pre-concentration technique, whereby an extra plug of 12.5% ammonium hydroxide was hydrodynamically injected at 20 mbar pressure for five seconds. This method was validated by combining 5 mM AMAc with 50 mM MeOH as a sheath solution with 500 mM FA at pH 2.0 as BGE, resulting in a significant increase in the separation of Ile and Leu (R s = 2.13) and high repeatability.…”

Capillary electromigration methods for food analysis and Foodomics: Advances and applications in the period March 2021 to March 2023

“…Negative pressure avoided a suction effect at the capillary tip that decreases separation efficiency during electrospray ionization (ESI) process. As shown in Figure 1, these optimized parameters along with the optimization of capillary voltage (4 kV), nebulizing gas pressure (10 psi), drying gas temperature (300 • C), and drying gas flow rate (10 L/min) of MS analysis allowed a selective analysis of AAs with reliable and reproducible results with a simple preparation of complex samples [27].…”

“…This method offers a rapid evaluation of the meat ripening process determined by CZE–UV due to requiring lower analysis times than traditional AA determinations, such as nonprotein nitrogen determination [26]. Besides, a CE–MS/MS method was developed and validated to analyze AAs from different complex samples for the quality control of food, sport supplements, and bioanalytical samples such as human plasma without a derivatization process [27]. In fact, only a dilution process of samples was carried out for the analysis of AAs (valine, isoleucine, and leucine) by CE–MS/MS excepting human plasma sample, which were treated with acetonitrile for the protein precipitation.…”