Development of a UHPLC-QQQ-MS/MS Method for Simultaneous Phytohormone Quantification and Tissue-Dependent Distribution in Loquat

Loquat (Eriobotrya japonica), an economically important evergreen fruit tree, exhibits an unusual reproductive cycle characterized by autumn–winter flowering and spring fruit maturation. However, the endogenous hormonal basis underlying this distinctive developmental pattern remains poorly understood, partly due to the lack of reliable multi-hormone analytical platforms for complex perennial fruit tissues. In this study, we established a robust targeted UHPLC-QQQ-MS/MS method for the simultaneous quantification of 11 endogenous phytohormones and related metabolites, including indole-3-acetic acid (IAA), abscisic acid (ABA), gibberellins (GA1, GA3, GA4, and GA7), and cytokinins (tZR, iPR, DHZ, iP, and zeatin). Sample preparation and chromatographic conditions were systematically optimized to improve extraction efficiency, to reduce matrix interference, and to maximize detection sensitivity. The method exhibited high linearity (R2 > 0.990), satisfactory recoveries (80.02–120.09%), low limits of detection, and high repeatability. Application of this platform to eight different loquat tissues revealed pronounced tissue-specific patterns of phytohormone distribution. Seeds contained exceptionally high levels of GA3 and zeatin, whereas stamens exhibited marked enrichment of IAA. Mature leaves showed relatively high ABA levels, whereas fruit flesh retained substantial cytokinin pools. These findings provide a reliable analytical workflow for phytohormone quantification in perennial fruit crops and establish a foundational dataset of tissue-specific hormone distribution in loquat, which may serve as a valuable reference for future studies on reproductive development and horticultural trait improvement.