Optimisation of ascorbate and glutathione assays for woody perennial bud tissues: overcoming matrix effects and storage instability
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ID: 322723
2026
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Abstract
Quantifying ascorbate (ASC) and glutathione (GSH) in woody perennial tissues presents significant methodological challenges due to complex tissue matrices and the lability of these redox-active metabolites. We adapted established enzymatic microplate assays for ASC and GSH quantification to mature dormant grapevine buds, a structure containing protective scales, hairs, and lignified components. Key optimisations included: (1) use of 1 M HClO₄ rather than 0.2 M HCl for effective enzyme inactivation and matrix disruption; (2) sample to solvent ratios of 1:10 to 1:15; and (3) extended centrifugation to effectively sediment organic components. Standard addition experiments revealed significant matrix effects, with recovery of approximately 80 % for ASC and 65 % for GSH. The optimised method successfully determined >90 % reduction status for both metabolites in mature buds. Critically, we observed severe ASC instability during frozen storage, with approximately 80 % loss of ASC after four months at -80 °C, while GSH remained stable. These findings underscore the necessity of immediate analysis for ASC quantification in woody tissues and provide a validated protocol for studying redox metabolism in perennial bud tissues.
| Reference Key |
openalex_W7171571377
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| Authors | Juwita Ratna Dewi, Pragya Poudel, Wenyi Xu, Santiago Signorelli, Michael Considine |
| Journal | Tree physiology |
| Year | 2026 |
| DOI |
10.1093/treephys/tpag103
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| URL | |
| Keywords | Keywords not found |
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