Determination of brassinosteroids in the sub‐femtomolar range using dansyl‐3‐aminophenylboronate derivatization and electrospray mass spectrometry

Svatoš, Aleš; Antonchick, Andrey P.; Schneider, Bernd

doi:10.1002/rcm.1413

Cited by 55 publications

(31 citation statements)

References 14 publications

(14 reference statements)

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“…Triply deuterated brassinosteroids, [26-2 H 3 ]3-epicastasterone, and [26-2 H 3 ]3-epibrassinolide [31] (0.4 µg in 10 µl of 75% EtOH each), were added as internal standards. Several solvent partitioning and chromatographic steps resulted in brassinosteroid-containing fractions, which after derivatization with an excess of dansyl-3-aminophenylboronic acid were analyzed by liquid chromatography (HPLC) with electrospray ionization (ESI) and using an established selected ion-monitoring mass-spectrometric (SIM-MS) detection (HPLC-ESI-SIM-MS) method [10,32]. Identification of brassinosteroids based on the relative retention time of 22,23-dansyl-3-aminophenylboronates (Table 1).…”

Section: Brassinosteroids In Arabidopsis Thaliana and Secale Cerealementioning

confidence: 99%

“…After a two-day incubation, the plant material was extracted according to the procedure used to detect endogenous brassinosteroids. Brassinosteroid-containing fractions were converted to their dansyl-3-aminophenylboronates and analyzed by HPLC-ESI-SIM-MS [10,32].…”

Section: Brassinosteroids In Arabidopsis Thaliana and Secale Cerealementioning

confidence: 99%

“…Based on the retention times of authentic standards (compound 1: R t 23.0 min; 2: R t 20.9 min; 3: R t 21.3 min; 4: R t 19.1 min), the brassinosteroid-containing fraction (R t 17.5 -24.5 min) was collected and evaporated. The residue was converted to bisdansyl-3-aminophenylboronates by being heated to 62 • C for 30 min with a solution of dansyl-3-aminophenylboronic acid (3 mg ml −1 ) in a mixture of pyridine and acetonitrile (1 : 19) and used for analysis by HPLC-ESI-SIM-MS as previously described [10,32].…”

Section: Isolation and Analysis Of Brassinosteroid Metabolitesmentioning

confidence: 99%

See 2 more Smart Citations

Reversible Conversion In The Brassinosteroid Quartet Castasterone, Brassinolide And Their 3β-Epimers

Antonchik¹,

Svatoš

Konstantinova³

et al. 2006

Zeitschrift Für Naturforschung B

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The metabolism of deuterated brassinosteroids has been studied in excised leaves of Secale cereale, and in vitro in seedlings of Arabidopsis thaliana and cell suspension cultures of Lycopersicon esculentum. In addition to the known biosynthetic conversion of castasterone to brassinolide and epimerization to 3-epicastasterone, inversion of the 3α-configured hydroxyl group of brassinolide to a 3β -configured one in 3-epibrassinolide has been observed using liquid chromatography (HPLC) with electrospray ionization (ESI) and selected ion-monitoring mass-spectrometry (SIM-MS). Administration of deuterated 3-epicastasterone and 3-epibrassinolide to Arabidopsis and Secale seedlings resulted in the formation of castasterone and brassinolide, respectively, indicating conversion of configuration at C-3 of brassinosteroids is reversible.

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Section: Brassinosteroids In Arabidopsis Thaliana and Secale Cerealementioning

confidence: 99%

Section: Brassinosteroids In Arabidopsis Thaliana and Secale Cerealementioning

confidence: 99%

Section: Isolation and Analysis Of Brassinosteroid Metabolitesmentioning

confidence: 99%

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Reversible Conversion In The Brassinosteroid Quartet Castasterone, Brassinolide And Their 3β-Epimers

Antonchik¹,

Svatoš

Konstantinova³

et al. 2006

Zeitschrift Für Naturforschung B

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“…Derivatization protocols References Diazomethane Sample was methylated using a stream of diazomethane gas [30] Sample was dissolved in 10% (v/v) methanol in diethylether and was methylated by reaction with diazomethane for 10 min at 0°C [7] Incubate at room temperature in methanol solution [81,90] creasing the volatility of plant hormones, so that the derived plant hormones are more suitable for GC/MS analysis. Although LC/MS can analyze plant hormones directly, derivatization is employed to improve ionization efficiency of the derived hormones, which can improve sensitivity by 10-to 1000-fold compared with underived hormones, and so has received increasing attention for the analysis of plant hormones [86][87][88].…”

Section: Chromatography and Chromatography/mass Spectrometrymentioning

confidence: 99%

Progress in quantitative analysis of plant hormones

Sun

Wang

et al. 2011

Chin. Sci. Bull.

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Plant hormones are small molecular natural products that regulate all plant developmental processes at low concentrations. Quantitative analysis of plant hormones is increasingly important for in-depth study of their biosynthesis, transport, metabolism and molecular regulatory mechanisms. Although plant hormone analysis remains a bottleneck in plant scientific research owing to the trace concentrations and complex components in plant crude extracts, much progress has been achieved in the development of extraction, purification and detection techniques in recent years. Solid phase extraction and chromatography/mass spectrometry have been applied widely for purification and quantitative analysis of plant hormones owing to their high selectivity and sensitivity. Purification methods such as liquid partition and immunoaffinity chromatography, and detection methods including immunoassay and electrochemical analysis, are employed. The advantages and disadvantages of these methods are discussed. In situ, real-time and multi-plant hormone profiling will comprise mainstream techniques for quantitative analyses in future studies on the regulatory mechanisms and crosstalk of plant hormones. defense responses are all triggered by plant hormones. A plant hormone, acting as a plant-development controller or a plant-environment mediator, can influence crop yield directly or indirectly, therefore research into plant hormone physiology has become an important target for agriculture development. However, it is noteworthy that endogenous level of each plant hormone is very low [1], and quantitative analysis of these compounds is still extremely difficult for both biologists and analytical chemists. It is a challenge for researchers to enrich these low-level natural compounds from plant extracts in which large amounts of interfering substances are present. In addition, the limited quantities of plant tissue available for hormone quantification is another challenge. Consequently, the analytic methods must be extremely selective and sensitive. The quantitative analysis of plant hormones typically includes the following key points: (i) the plant materials must

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“…Yamashita et al reported a sensitive and specific assay method for estrogen o-quinones using LC/ESI-MS/MS combined with the o-phenylenediamine-derivatization. 7) Brassinosteroids (vicinal diol-containing plant hormones) were also analyzed by a derivatization followed by LC/ESI-MS. 8) However, to the best of our knowledge, a practical derivatization procedure specific for the bioanalysis of steroids having a vicinal diol has not been reported.…”

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confidence: 99%

LC/MS/MS of Steroids Having Vicinal Diol as Electrospray-Active Boronates

Higashi

Kawasaki

Matsumoto

et al. 2013

CHEMICAL & PHARMACEUTICAL BULLETIN

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A derivatization procedure with (3-dimethylaminophenyl)dihydroxyborane (DAPB) was introduced to enhance the detectability of steroids having a vicinal diol in LC/electrospray ionization (ESI)-MS/MS. DAPB reacted with the vicinal diol on the steroids [4β-hydroxycholesterol (4-HCh), pregnanetriol (PT) and 20R,22R-dihydroxycholesterol] in pyridine at 50°C within 1 h. The resulting DAPB-derivatives were highly responsive in ESI-MS operating in the positive-ion mode and gave characteristic product ions during MS/ MS, which enabled sensitive detection using a selected reaction monitoring mode; the detection responses of the DAPB-derivatives were increased by 20-160-fold over those of the intact steroids and the limits of detection were in the low femtomole or attomole range. The derivatization procedure was successfully applied to biological sample analysis; the derivatization followed by LC/ESI-MS/MS enabled the specific detection of trace amounts of 4-HCh in human plasma and PT in human urine with a small sample volume, simple pretreatment and short chromatographic run time.Key words derivatization; electrospray ionization-MS/MS; steroid; vicinal diol; detectability A specific and sensitive method for the characterization and determination of endogenous steroids in biological fluids and tissues is useful for the elucidation of the nature, diagnosis and treatment of diseases. LC coupled with electrospray ionization (ESI)-MS/MS is widely used for steroid analysis due to its specificity, versatility and simultaneous multi-analyte quantification capability.1-3) However, not all steroids are suitable for ESI-MS/MS. One of the limitations in ESI-MS/MS for some steroids is the poor ionization and fragmentation behavior, leading to low sensitivity; current MS/MS instruments are extremely sensitive, but still their absolute sensitivity is analyte-dependent. To overcome this limitation, derivatization of the steroids has often been employed in LC/ESI-MS/ MS [4][5][6] ; a suitable derivatization can enhance the ionization efficiency of a poorly ionizable steroid in ESI-MS and assist fragmentation suitable for the selected reaction monitoring (SRM) mode.In most of the derivatization procedures reported until now, an ESI-active (proton-affinitive or permanently-charged) moiety is introduced to the target steroid through a hydroxyor an oxo-group. [4][5][6] These procedures have been proven to be very useful for LC/ESI-MS/MS of some steroids, but the derivatization utilizing a vicinal bifunctional group, such as vicinal diol and vicinal diketone, has not been fully examined. By such a derivatization, more specific detection and quantification of the target steroid may become possible. Yamashita et al. reported a sensitive and specific assay method for estrogen o-quinones using LC/ESI-MS/MS combined with the o-phenylenediamine-derivatization.7) Brassinosteroids (vicinal diol-containing plant hormones) were also analyzed by a derivatization followed by LC/ESI-MS. 8) However, to the best of our knowledge, a practical derivatization p...

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Determination of brassinosteroids in the sub‐femtomolar range using dansyl‐3‐aminophenylboronate derivatization and electrospray mass spectrometry

Cited by 55 publications

References 14 publications

Reversible Conversion In The Brassinosteroid Quartet Castasterone, Brassinolide And Their 3β-Epimers

Reversible Conversion In The Brassinosteroid Quartet Castasterone, Brassinolide And Their 3β-Epimers

Progress in quantitative analysis of plant hormones

LC/MS/MS of Steroids Having Vicinal Diol as Electrospray-Active Boronates

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