Plant Hormone Analysis Service | Targeted Phytohormone Profiling & Quantification Panel

Plant Hormone Signaling & Targeted Quantification

Plant hormones (phytohormones) are a structurally diverse set of signaling molecules that orchestrate virtually every aspect of plant life—from seed germination, root architecture, and shoot development to flowering, fruit ripening, and stress adaptation. These compounds operate at trace concentrations (pmol–fmol per gram fresh weight) through complex synergistic and antagonistic networks, making their accurate quantification both critically important and analytically challenging.

The major hormone families—auxins, cytokinins (CKs), gibberellins (GAs), abscisic acid (ABA), ethylene (ET), jasmonates (JAs), salicylic acid (SA), brassinosteroids (BRs), and strigolactones (SLs)—each regulate distinct but interwoven processes. For example, the auxin–cytokinin balance determines shoot vs. root architecture, ABA drives stomatal closure under drought, and the JA/ET module orchestrates defense against necrotrophic pathogens.

Targeted phytohormone panel metabolomics via LC–MS/MS and GC–MS/MS provides the most reliable approach for accurate hormone quantification at the pathway level. Unlike untargeted approaches, which survey all detectable signals and may miss low-abundance regulators, targeted MRM-based methods achieve the attomole-level sensitivity and structural specificity required for trace hormones. This enables you to:

(i) measure absolute hormone pool sizes and key metabolic ratios (e.g., IAA/CK, ABA/JA, GA/BR) that define developmental and stress states

(ii) monitor specific regulatory nodes such as auxin signaling, ABA/JA/ET stress cascades, GA/BR growth regulation, strigolactone/karrikin branching control, and peptide hormone signaling pathways

(iii) compare across plant tissues, developmental stages, stress treatments, and genetically modified lines with batch-to-batch data comparability.

Key Challenges in Targeted Phytohormone Analysis

Phytohormone analysis presents a unique set of analytical challenges not encountered in routine metabolomics. Low endogenous concentrations, severe matrix interference from pigments and polyphenols, and the need to cover multiple chemically distinct classes in a single run all contribute to the difficulty. Creative Proteomics addresses each barrier with targeted solutions:

• Low abundance across structurally diverse classes: Phytohormones span a wide polarity range—from hydrophobic BRs to highly polar CK nucleotides—and exist at sub-ppm levels. Our workflow combines class-specific SPE enrichment and multi-mode LC (RP–C18 for most classes; HILIC for polar CKs and precursors) to maximize recovery across the full chemical spectrum. MRM acquisition on triple-quadrupole platforms (SCIEX QTRAP 6500+, Agilent 6495C) delivers attomole-level LODs.
• Matrix-driven ion suppression: Pigmented leaf tissue, resinous seed coats, and woody roots introduce co-eluting compounds that suppress ionization. We apply two-step SPE cleanup (mixed-mode anion exchange + RP) and matrix-matched calibration to correct for suppression, plus up to 30 isotope-labeled internal standards (D6-ABA, D5-tZ, D2-IAA, D6-SA, D2-JA, D2-GA4, D3-DHZ, and others) for per-analyte normalization.
• Isomer and epimer resolution: Many hormone classes contain structural and stereoisomers with distinct biological activities—cis- vs. trans-zeatin, GA1 vs. GA3, D/L-amino acid conjugates. We deploy chiral LC phases and optimized gradient programs to achieve baseline separation of critical isomers, supported by authenticated standard libraries for each target.
• Cross-class coverage in a single analytical run: Comprehensive hormonomics requires simultaneous quantification of compounds with vastly different physicochemical properties. Our integrated method uses time-segmented MRM scheduling, polarity switching, and derivatization strategies (dansyl chloride for BRs; MSTFA for GAs by GC–MS) to capture 100+ analytes from 10+ hormone classes in a single injection sequence.
• Labile compound preservation: GAs, BRs, and certain CK phosphates are prone to degradation during extraction and storage. We employ flash-freeze harvest protocols, cold methanolic extraction with antioxidant stabilization (BHT, ascorbic acid), and minimized sample handling to preserve labile targets.

Phytohormone Detection Panels & Pathway Coverage

Creative Proteomics offers a suite of LC–MS/MS and GC–MS/MS-based targeted panels covering major phytohormone classes, pathway-specific intermediates, and signaling metabolites. Panels are pre-configured for rapid deployment or fully customizable to match your species, tissue, and research question.

Standard Quantification Panels

Pathway-Focused Panels

Functional Biomarker Panels

Customization & Add-ons

Why Choose Our Phytohormone Analysis Service?

• Curated Library of 150+ Phytohormones & Signaling Metabolites
  Our in-house MS/MS library covers auxins, CKs, GAs, ABA, BRs, JAs, SA, SLs, peptide hormones, oxylipins, and related conjugates—enabling rapid panel configuration, confident annotation, and seamless expansion across interconnected pathways.
• Up to 30 Isotope-Labeled Internal Standards
  We deploy hormone-specific stable isotope-labeled internal standards (D6-ABA, D5-tZ, D2-IAA, D6-SA, D2-JA, D2-GA4, D3-DHZ, 13C6-IAA, and others) to correct for matrix effects, extraction recovery, and ionization efficiency—ensuring absolute quantification accuracy across structurally diverse targets.
• Advanced Matrix Cleanup for Complex Tissues
  Data undergo two-step SPE cleanup, isobaric interference correction, ion ratio QC, and manual peak review by experienced mass spectrometrists—ensuring data integrity even in challenging matrices such as pigmented leaf tissue, woody roots, resinous seed coats, and senescent material.
• Batch-Reproducible & Method-Locked Workflows
  Ideal for multi-batch studies, long-term developmental time courses, and multi-site comparisons. Methods are locked per project; inter-batch bridging samples and system suitability benchmarks ensure data comparability across weeks or months of acquisition.

LC–MS/MS Platform & Method Performance

Internal Standards & Calibration

• Internal Standards: Up to 30 stable isotope-labeled phytohormones and pathway-specific analogs
• Calibration Strategy: 6–8 point standard curves, matrix-matched or surrogate matrix as appropriate
• System Suitability: Ion ratio QC, retention time window verification, peak shape assessment, carryover checks, and blank subtraction per batch

Phytohormone Analysis Workflow

1

Panel Design & Method Configuration

We align panel composition with your hormone classes of interest (e.g., auxin signaling, ABA/JA stress, GA regulation, BR/sL profiling), sample matrix, and sensitivity targets. Internal standards, derivatization strategy, and LC/GC conditions are selected and documented in a project-specific method statement.

2

Sample Preparation & Stabilization

Samples are processed using matrix-specific protocols: flash-freeze in liquid N₂ within 30 seconds of harvest, cryogenic grinding, and cold solvent extraction with antioxidant stabilization. Labile GAs, BRs, and peptide hormones receive dedicated handling protocols. Each batch includes process blanks and preparation controls.

3

LC–MS/MS or GC–MS/MS Acquisition

Quantification is performed using MRM-based targeted methods under validated acquisition settings. Polar/thermolabile hormones (auxins, CKs, ABA, JAs, BRs) are analyzed by LC–MS/MS; volatile/thermostable hormones (GAs, ACC) are analyzed by GC–MS/MS. All runs include system suitability tests and internal standard monitoring.

4

Quantification & Quality Review

Hormone concentrations are calculated via 6–8 point calibration curves and normalized to isotope-labeled internal standards. Data are reviewed for linearity (R² ≥ 0.99), precision (CV ≤ 10%), ion ratio conformity, and carryover. QC replicates are embedded at 10% frequency per batch.

5

Data Delivery & Biological Context

You receive a complete data package including raw and processed data, QC and calibration reports, and annotated results with KEGG/HMDB/CAS identifiers. Pathway maps and fold-change visualizations are provided for hormone signaling networks to aid biological interpretation.

Sample Submission Guide for Phytohormone Analysis

Notes

• Avoid repeated freeze-thaw cycles—plant hormones are highly labile
• Collect all samples at the same time of day (circadian hormone variation can exceed 2-fold for IAA and ABA)
• Clearly label each tube with sample ID, tissue type, developmental stage, date and time of collection
• Submit sample list (Excel preferred) with matching metadata including treatment conditions
• Contact us before sending rare, low-volume, or highly pigmented matrices for protocol optimization

Deliverables: What You Receive from Phytohormone Analysis

• Quantification tables: absolute (pmol/g FW) and/or relative hormone concentrations (.csv / Excel)
• Calibration & QC report: calibration curves, accuracy, precision, recovery, carryover checks per hormone class
• Raw & processed data: instrument raw files, processed peak lists, MRM chromatograms
• Method documentation: LC–MS/MS or GC–MS/MS parameters, derivatization protocol, internal standard setup
• Pathway annotations: KEGG / HMDB / CAS IDs linked to quantified phytohormones
• Final summary report: sample notes, QC results, interpretation highlights

Representative MRM chromatogram showing baseline-separated phytohormone peaks (IAA, ABA, JA, tZ, GA4) under optimized LC–MS/MS conditions.

Calibration curve of IAA across 8 concentration levels, with linear regression (R² = 0.999) and indicated LOD/LOQ thresholds.

Batch-wise overlay of ABA concentrations from five QC samples, demonstrating intra- and inter-batch reproducibility with low variance.

Integrated phytohormone pathway network illustrating quantified analytes from auxin, CK, GA, ABA, JA, and BR pathways, colored by fold-change.

Applications of Phytohormone Profiling

Our phytohormone profiling service supports researchers and breeders across plant science disciplines—from model species to crop plants and medicinal herbs. Applications span fundamental developmental biology, stress physiology, agricultural biotechnology, and multi-omics integration.

Integrated Transcriptome and Hormone Profiling Reveals Dynamic Responses to Moderate Dehydration Stress in Arabidopsis

Background

Plant responses to dehydration stress involve complex molecular and hormonal changes coordinated across multiple timescales. While the role of ABA in drought response is well established, the temporal dynamics of the full hormone network—including JA, SA, IAA, CKs, and GAs—during moderate dehydration remained poorly characterized. Single-hormone measurements or single time-point analyses fail to capture the regulatory crosstalk that drives adaptive responses.

Challenge: Characterize the temporal dynamics of genome-wide gene expression and comprehensive hormone profiles during moderate dehydration stress in Arabidopsis, with a focus on hormone–transcriptome integration to identify regulatory hubs and crosstalk mechanisms.

Key Findings (from the published study)

• Bi-phasic ABA accumulation observed during moderate dehydration: an early rapid increase (stomatal closure phase, 1–6 h) followed by sustained accumulation (protective gene induction phase, 12–72 h)
• Dynamic transcriptome reprogramming with distinct early (0–6 h) and late (12–72 h) response phases, each characterized by unique co-regulated gene sets
• Coordinated JA–ABA signaling: Dehydration-induced JA accumulation preceded SA changes, and JA-responsive genes showed significant overlap with ABA-responsive transcripts, revealing intimate crosstalk between stress signaling pathways
• AP2/ERF transcription factors identified as early dehydration response regulators, linking hormone signaling to transcriptional reprogramming
• Touch stress-responsive genes distinguished from dehydration-specific responses through the time-resolved experimental design

Process Insight

This study demonstrates that single-hormone (ABA-only) measurements would have revealed the biphasic pattern but missed the JA–ABA crosstalk and the differential regulation of auxin, CK, and GA pathways associated with growth suppression under stress. Comprehensive multi-hormone profiling using LC-ESI-MS/MS with isotope-labeled internal standards was essential to build a systems-level understanding of dehydration responses—an insight directly applicable to engineering drought-tolerant crops.

Where Our Phytohormone Panel Fits

While this study used a research-grade hormone profiling approach, Creative Proteomics supports similar plant stress research by providing:

• Comprehensive phytohormone panels covering auxins, CKs, GAs, ABA, JAs, SA, ACC, BRs, and SLs—all from a single sample submission
• LC–MS/MS and GC–MS/MS (MRM) with isotope-dilution for absolute quantification and cross-batch comparability across multi-timepoint studies
• Pathway-linked outputs (KEGG IDs) and optional multi-omics integration (untargeted metabolomics, transcriptomics, proteomics) for mechanistic insights into hormone-regulated networks

Reference

1. Urano, K., Maruyama, K., Jikumaru, Y., Kamiya, Y., Yamaguchi-Shinozaki, K., & Shinozaki, K. (2017). Analysis of plant hormone profiles in response to moderate dehydration stress. The Plant Journal, 90(1), 17–36. https://doi.org/10.1111/tpj.13472