Corrections for interferences and extraction conditions make a difference: use of the TBARS assay for lipid peroxidation of orthodox Spartina pectinata and recalcitrant Spartina alterniflora seeds during desiccation

摘要

Lipid peroxidation and membrane damage are often proposed as causes of recalcitrant seed death, and the thiobarbituric acid reactive substances (TBARS) assay is commonly used to measure lipid peroxidation. However, several artefacts can cause an overestimation of TBARS values, and these have not been routinely addressed in experiments with recalcitrant seeds. In the present report, TBARS was assayed as recalcitrant Spartina alterniflora and orthodox S. pectinata seeds were dried rapidly. Using the traditional Heath and Packer (1968) protocol with tissue extraction at 4 degrees C, S. alterniflora had higher overall TBARS values than S. pectinata, and TBARS products increased when recalcitrant S. alterniflora and orthodox S. pectinata seeds were dried. However, when corrections for interfering substances, such as sugars and anthocyanins, were made, the TBARS values between the two species were almost identical. When seeds were freeze-clamped in liquid nitrogen prior to extraction, TBARS did not increase during desiccation for either species. These findings may indicate that lipid peroxidation is not the cause of desiccation-induced death in S. alterniflora. Therefore, freeze-clamping during tissue extraction and corrections for TBARS interfering substances must be applied to avoid overestimation of lipid peroxidation values.