Volatile fingerprints of seeds of four species indicate the involvement of alcoholic fermentation, lipid peroxidation, and Maillard reactions in seed deterioration during ageing and desiccation stress

Abstract

The volatile compounds released by orthodox (desiccation-tolerant) seeds during ageing can be analysed using gas chromatography-mass spectrometry (GC-MS). Comparison of three legume species (Pisum sativum, Lathyrus pratensis, and Cytisus scoparius) during artificial ageing at 60% relative humidity and 50 °C revealed variation in the seed volatile fingerprint between species, although in all species the overall volatile concentration increased with storage period, and changes could be detected prior to the onset of viability loss. The volatile compounds are proposed to derive from three main sources: alcoholic fermentation, lipid peroxidation, and Maillard reactions. Lipid peroxidation was confirmed in P. sativum seeds through analysis of malondialdehyde and 4-hydroxynonenal. Volatile production by ageing orthodox seeds was compared with that of recalcitrant (desiccation-sensitive) seeds of Quercus robur during desiccation. Many of the volatiles were common to both ageing orthodox seeds and desiccating recalcitrant seeds, with alcoholic fermentation forming the major source of volatiles. Finally, comparison was made between two methods of analysis; the first used a Tenax adsorbent to trap volatiles, whilst the second used solid phase microextraction to extract volatiles from the headspace of vials containing powdered seeds. Solid phase microextraction was found to be more sensitive, detecting a far greater number of compounds. Seed volatile analysis provides a non-invasive means of characterizing the processes involved in seed deterioration, and potentially identifying volatile marker compounds for the diagnosis of seed viability loss.

Fig. 1.

Ageing of Pisum sativum, Lathyrus pratensis, and Cytisus scoparius seeds, and desiccation of Quercus robur seeds caused loss of viability, shown by a decline in total germination and a reduction in seed vigour, indicated by an increase in the amount of time taken to complete germination. The graphs show the total germination of P. sativum (A) seeds following ageing for 0 (white circles), 8 (grey circles), 12 (black circles), 15 (white triangles), 25 (grey triangles), and 55 d (black triangles); C. scoparius (B) seeds aged for 0 (white circles), 21 (white triangles), 49 (grey triangles). and 63 d (black triangles); L. pratensis (C) seeds aged for 0 (white circles), 35 (grey triangles), and 70 d (black triangles); and Q. robur (D) seeds desiccated for 0 (white circles), 3 (white triangles), 6 (grey triangles), and 13 d (black triangles). Each data point represents the mean ±SE of four (C. scoparius and L. pratensis) or five (P. sativum and Q. robur) replicates. Letters indicate statistically significant differences (P < 0.05) in total germination between ageing/desiccation treatments.

Fig. 2.

The amount of volatile compounds released by the seeds increased with the ageing/desiccation period. The stacked bars represent the overall concentration of volatiles and volatile composition (indicated by different shading patterns) released by Pisum sativum (A), Cytisus scoparius (B), and Lathyrus pratensis (C) seeds during artificial ageing, and Quercus robur acorns during desiccation (D). Volatile compounds were trapped using Tenax and analysed using GC-MS. The lines and open circles represent the mean total germination at each ageing/desiccation time point. Each bar and data point represents the mean ±SE of five replicates for P. sativum and Q. robur, and four for C. scoparius and L. pratensis. The stacked bars corresponding to ‘seed’ on the x-axis of the Q. robur graph (D) represent the volatiles released after removal of the seed coat from Q. robur seeds that had been desiccated for 13 d. (This figure is available in colour at JXB online.)

Fig. 3.

The volatile compounds present in the headspace of desiccated Quercus robur acorns and aged Pisum sativum, Lathyrus pratensis, and Cytisus scoparius seeds were analysed using solid phase microextraction (SPME). The seed material had been freeze-dried, ground, and stored at –75 °C prior to analysis. The stacked columns represent the total volatile concentration and composition (indicated by different shading patterns) released by Q. robur acorns following desiccation (A, E, I, M), and P. sativum (B, F, J, N), L. pratensis (C, G, K, O), and C. scoparius (D, H, L, P) seeds following artificial ageing. The overall volatile composition shows the four major volatiles: ethanol, methanol, acetaldehyde, and acetone, along with the sum of all other volatiles (A–D). The other volatiles are divided into groups: alcohols (E–H); aldehydes (I–L); and miscellaneous compounds (M–P). Each bar represents the mean ±SE of three replicates. Plots with the y-axis shown on the left share the same scale as the left-hand plot within the same row, whilst plots with the y-axis on the right have the same scale as the right-hand plot within the same row. Note that the scale of plot K is multiplied by 10. (This figure is available in colour at JXB online.)