Phytohormones as Important Biologically Active Molecules – Their Simple Simultaneous Detection

Abstract

Phytohormones, their functions, synthesis and effects, are of great interest. To study them in plant tissues accurate and sensitive methods are required. In the present study we aimed at optimizing experimental conditions to separate and determine not only plant hormones but also their metabolites, by liquid chromatography coupled with a UV-VIS detector. The mixture we analyzed was composed of benzyladenine, kinetin, trans-zeatin, cis-zeatin, dihydrozeatin, meta-topolin, ortho-topolin, α-naphthalene acetic acid, indole-3-acetic acid, trans-zeatin-7-glucoside, trans-zeatin-O-glucoside, trans-zeatin-9-riboside, meta-topolin-9-riboside and ortho-topolin-9-riboside. We measured the calibration dependences and estimated limits of detection and quantification under the optimal chromatographic conditions (column: Polaris C18; mobile phase: gradient starting at 2:98 (methanol:0.001% TFA) and was increasing to 55:45 during twenty minutes, and then decreasing for 10 min to 35:65, flow rate: 200 µL·min-1, temperature: 50 °C, wavelength: 210 nm). The detection limits for the target molecules were estimated as tens of ng per mL. We also studied the effect of flax extracts on the phytohormones’ signals. Recovery of aliphatic and aromatic cytokinins, metabolites of cytokinins and auxins were within the range from 87 to 105 %. The experimental conditions were tested on a mass selective detector. In addition we analysed a commercial product used for stimulation of roots formation in cuttings of poorly rooting plants. The determined content of α-naphthalene acetic acid was in good agreement with that declared by the manufacturer.

A Lobularia maritima LmSAP protein modulates gibberellic acid homeostasis via its A20 domain under abiotic stress conditions

“…Rescue of dwarf phenotypes in LmSAPΔA20 and LmSAPΔA20-ΔAN1overexpressing tobacco lines under abiotic stress conditions by application of exogenous GA 3 The overexpression of LmSAPΔA20 and LmSAPΔA20-ΔAN1 results in dwarfism and smaller leaves in tobacco plants exposed to salt or osmotic stress under in vitro conditions [19]. To test whether exogenous GA 3 treatment could rescue this phenotype, NT tobacco plants and two homozygous transgenic plants from LmSAPΔA20 and LmSAPΔA20-ΔAN1 constructs (labeled C2, C6, D2, and D9) were treated with 100 μM GA 3 on MS solid medium supplemented with 300 mM NaCl or 300 mM mannitol.…”

“…Perception of stressful environmental conditions induces a specific stress hormone signature specifying a proper response, which is efficient in terms of fitness. Several plant growth regulators, such as phytohormones, regulate many physiological and metabolic processes that help plants to adapt to environmental stresses [3,4]. The phytohormone gibberellin (GA), also known as gibberellic acid [5], regulates various crucial growth and developmental processes, such as seed germination, stem elongation, leaf expansion, and reproductive development, in most plant species [6].…”

Potentiometric and Voltammetric Study of 6‐Benzylaminopurine and Its Derivatives

“…The study of interaction between benzylaminopurines and metal ions such as Cu, Co, Fe, Pt, and Pd showed that the BAP derivative is able to create dinuclear complexes in its protonated form 5–7. To analyze adenine‐type cytokinins, attention was paid to chromatography (HPLC/MS, HPLC/ED GC/MS), immunochemical methods (ELISA, RIA) 2,8,9 and electrochemical methods 2,10. From the electroanalytical point of view adenine and its derivatives, including 6‐BAP, represent a group of substances with typical redox properties.…”

Classification and characterization of unknown cytokinins into essential types by in‐source collision‐induced dissociation electrospray ionization ion trap mass spectrometry

“…Two common methods for qualification and quantification of cytokinins are mass spectrometry (MS) and immunoassays. Because of its high sensitivity and selectivity, MS coupled with liquid chromatography (LC) has become increasingly popular . Although the effective range and the sensitivity of the immunoassay are similar to those of LC/MS, there are some problems with immunoassays, including cross‐reactivity, imperfect validation and variable results for analyses of plant sample .…”

“…Because of its high sensitivity and selectivity, MS coupled with liquid chromatography (LC) has become increasingly popular. [11][12][13][14] Although the effective range and the sensitivity of the immunoassay are similar to those of LC/MS, there are some problems with immunoassays, including cross-reactivity, imperfect validation and variable results for analyses of plant sample. [15] Using MS, it is possible to identify a compound by simple interpretation of the mass spectrum; however, a more thorough analysis includes comparison of the obtained mass spectrum with that of a standard.…”