Evaluating methods for extraction of α-cellulose from leaves of Melaleuca quinquenervia for stable carbon and oxygen isotope analysis

Rationale: Purification of α-cellulose from plant tissues is commonly conducted to facilitate the reliable measurement of stable isotope ratios. Prior research has shown that different plant species and tissues react differently to standardised cellulose extraction techniques. Thus, no single method can be applied to all materials and careful consideration must be undertaken when selecting an extraction technique. 

Methods: In order to evaluate their suitability for use on Melaleuca quinquenervia leaves, a suite of eight different cellulose extraction techniques were tested. Leaves of this species are preserved in perched lakes on southeast Queensland's sand islands and are a focus of ongoing palaeoclimate research. Elemental analyser/isotope ratio mass spectrometry was used to measure stable carbon and oxygen isotopic ratios and sample composition was measured using Fourier transform infrared spectroscopy. 

Results: We demonstrate that the standard Brendel extraction technique, particularly with a higher reagent volume and longer boiling time, produces cellulose with the lowest spread in isotopic ratios among replicates, and with the fewest impurities detected by Fourier transform infrared spectroscopy. We also show that pre-treating the leaves to extract leaf wax lipids in order to enable paired analysis from the same sub-fossil leaves does not significantly affect the quality of the isotopic results. 

Conclusions: The standard Brendel method allows the most precise stable carbon and oxygen information to be retrieved from the leaves of M. quinquenervia. This unlocks the potential to study palaeoclimate proxy records from our study site and potentially throughout the natural range of the species across eastern Australia, Papua New Guinea and New Caledonia.