Abstract
Simple and efficient microwave-assisted acid hydrolysis (MAAH) of proteins was used for rapid quantification of α-aminoadipic semialdehyde (AAS) and γ-glutamic semialdehyde (GGS) as major protein oxidation markers. The precursor amino acid residues corresponding to AAS and GGS in oxidized proteins were derivatized by reductive amination with sodium cyanoborohydride (NaCNBH3) and p-aminobenzoic acid (ABA) followed by MAAH to generate the marker derivatives AAS–ABA and GGS–ABA. The quantification was performed using electrospray ionization liquid chromatography–mass spectrometry (ESI LC–MS). The important parameters for hydrolysis were optimized, which include the temperature, the reaction time, the acid concentration and volume as well as the microwave power. Compared to the conventional acid hydrolysis of 18–24h using 6–12M HCl at 110°C applied commonly in the literature and also in this work, MAAH of proteins can be completed as fast as in only 2–10min and, additionally, with a 3–5 times higher yield of the final derivatization products. Furthermore, a better agreement between the ratio of the detected derivatization products and the theoretical yields from the studied protein has also been achieved, which indicates that MAAH may serve as a more reliable method of acid hydrolysis for this purpose than that with conventional thermal heating. The MAAH method is demonstrated to be a time-saving, reproducible and efficient technique for studying AAS and GGS as protein oxidation markers using LC–MS.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.