Standardized protocols and procedures can precisely and accurately quantify non-structural carbohydrates

Abstract

Non-structural carbohydrates (NSCs), the stored products of photosynthesis, building blocks for growth and fuel for respiration, are central to plant metabolism, but their measurement is challenging. Differences in methods and procedures among laboratories can cause results to vary widely, limiting our ability to integrate and generalize patterns in plant carbon balance among studies. A recent assessment found that NSC concentrations measured for a common set of samples can vary by an order of magnitude, but sources for this variability were unclear. We measured a common set of nine plant material types, and two synthetic samples with known NSC concentrations, using a common protocol for sugar extraction and starch digestion, and three different sugar quantification methods (ion chromatography, enzyme, acid) in six laboratories. We also tested how sample handling, extraction solvent and centralizing parts of the procedure in one laboratory affected results. Non-structural carbohydrate concentrations measured for synthetic samples were within about 11.5% of known values for all three methods. However, differences among quantification methods were the largest source of variation in NSC measurements for natural plant samples because the three methods quantify different NSCs. The enzyme method quantified only glucose, fructose and sucrose, with ion chromatography we additionally quantified galactose, while the acid method quantified a large range of mono- and oligosaccharides. For some natural samples, sugars quantified with the acid method were two to five times higher than with other methods, demonstrating that trees allocate carbon to a range of sugar molecules. Sample handling had little effect on measurements, while ethanol sugar extraction improved accuracy over water extraction. Our results demonstrate that reasonable accuracy of NSC measurements can be achieved when different methods are used, as long as protocols are robust and standardized. Thus, we provide detailed protocols for the extraction, digestion and quantification of NSCs in plant samples, which should improve the comparability of NSC measurements among laboratories.

Browsing and fire decreases dominance of a resprouting shrub in woody encroached grassland

“…We combined lateral and fine root tissues from each shrub island and placed the roots in a cooler. All roots were washed in distilled water to remove all soil particles and then microwaved for 90 s to halt enzymatic activity (Landhäusser et al ). After microwaving, roots were oven‐dried at 65°C for 72 h. All root samples were milled to 40 mesh (400 µm) prior to ball milling (Wig‐L‐Bug, Henry Schein, Inc., Melville, NY, USA) and stored in glass vials in a −20°C freezer until NSC analysis.…”

The Possible Role of Non-Structural Carbohydrates in the Regulation of Tree Hydraulics

“…Indeed, the current model of embolism refilling has been extensively tested only in some species of the genus Populus (see [42] for details), but it should be validated in a broader range of species. Routinely adding NSC measurements to hydraulic measurements, by using standardized protocols for NSC sampling and analysis [126], could shed light onto possible connections between the hydraulic and carbohydrate components. Moreover, given the possible diel variations in NSC contents in different organs and positions in the trees [127], the sampling time for NSC analysis is also crucial for comparisons between studies.…”

Elevated temperature differently affects growth, photosynthetic capacity, nutrient absorption and leaf ultrastructure of Abies faxoniana and Picea purpurea under intra- and interspecific competition

“…The extraction was utilized for soluble sugar determination, and the residue for starch determination. To prepare the residue for starch determination, tubes with residue were left uncovered under the fume hood over the night to evaporate the residual ethanol (Landhäusser et al 2018). Glucose equivalents were used to assay starch and soluble sugar concentrations by the anthrone-sulfuric acid method (Yemm and Willis 1954).…”