0476577 - ÚEB 2018 RIV NL eng J - Journal Article
Oklešťková, Jana - Tarkowská, Danuše - Eyer, L. - Elbert, Tomáš - Marek, Aleš - Smržová, Z. - Novák, Ondřej - Fránek, M. - Zhabinskii, V.N. - Strnad, Miroslav
Immunoaffinity chromatography combined with tandem mass spectrometry: A new tool for the selective capture and analysis of brassinosteroid plant hormones.
Talanta. Roč. 170, AUG 1 (2017), s. 432-440. ISSN 0039-9140. E-ISSN 1873-3573
R&D Projects: GA MŠMT(CZ) LO1204; GA ČR GA14-34792S; GA ČR GJ15-08202Y
Institutional support: RVO:61389030 ; RVO:61388963
Keywords : Brassica napus * Brassinosteroids * Enzyme immunoassay * Immunoaffinity chromatography * Liquid chromatography-tandem mass spectrometry * Monoclonal antibodies
OECD category: Analytical chemistry; Biochemical research methods (UOCHB-X)
Impact factor: 4.244, year: 2017

Brassinosteroids (BRs) are plant-specific steroid hormones that play essential roles in the regulation of many important physiological processes in plant life. Their extremely low concentrations (~pmoles/g FW) in plant tissue and huge differences in polarity of individual members within the BR family hamper their detection and quantification. To address this problem, an immunoaffinity sorbent with broad specificity and high capacity for different BR metabolites containing a monoclonal antibody (mAb) against a BR spacer (20S)−2α,3α-dihydroxy-7-oxa-7α-homo-5α-pregnane-6-one-20 carboxylic acid (BR4812) was used for the rapid and highly selective isolation of endogenous BRs containing a 2α,3α-diol in ring A from minute plant samples. This enrichment procedure was successfully applied as a sample preparation method prior to quantitative analysis of BRs in real plant tissues by ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Use of immunoaffinity chromatography (IAC) increased the sensitivity of the UHPLC-MS/MS analysis owing to improvements in the BR signal-to-noise ratio (S/N) and matrix factor (MF). Although MF values of BRs analyzed in classical samples ranged from 8.9% to 47.4%, MF values for the IAC purified samples reached 44.5–96.6%. Thus, the developed IAC-UHPLC-MS/MS approach was shown to be a simple, robust, effective and extremely fast procedure requiring minute amounts of plant samples suitable for the quantitative profiling of many BR metabolites, helping to overcome the major problems associated with their determination in very complex plant matrices.
Permanent Link: http://hdl.handle.net/11104/0273051