Determination of Total Soluble Sugars in Pteridophytes Using the Anthrone Method.

Sugars serve as crucial integrators of both internal and environmental signals in plants, shaping the regulation of diverse physiological processes that occur throughout the plant's life span, from early embryogenesis to later senescence stages. During evolution, plants have developed various strategies to cope with abiotic stress. For example, the accumulation of osmolytes such as soluble sugars, which help protect against oxidative stress, stabilizes cellular membranes and preserves enzymes in the dry state. Precise quantification of total sugars is therefore essential for elucidating the biochemical and physiological strategies of plants in response to different conditions. Here, we present a detailed protocol for extracting and quantifying total soluble sugars in pteridophytes (lycophytes and ferns) from small amounts of leaf tissue (10 mg fresh tissue) using the anthrone method, a colorimetric assay in which carbohydrates react with the anthrone reagent under acidic conditions to form a blue-green complex whose concentration is measurable by spectrophotometry. The procedure was adapted for small sample volumes, incorporating ethanol extraction, preparation of a glucose standard curve, and absorbance measurement at 620 nm in 96-well plates. Quantification of total sugars in pteridophytes is essential for understanding the changes in metabolic responses. Likewise, using small amounts of plant tissue optimizes sugar extraction in plants with low biomass and minimizes impact on plant populations. © 2026 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Extraction of fresh plant tissue Basic Protocol 2: Reaction with anthrone and measurement Support Protocol 1: Preparation of anthrone reagent Support Protocol 2: Preparation of the glucose standard curve Basic Protocol 3: Calculation of total sugar concentration.