Dried plant specimens stored in herbaria are an untapped treasure chest of information on environmental conditions, plant evolution and change over many hundreds of years. Due to their delicate nature and irreplaceability, there is limited access for analysis to these sensitive samples, particularly where chemical data is obtained using destructive techniques. Fourier transform infrared spectroscopy (FTIR) is a chemical analysis technique that can be applied non-destructively to understand chemical bonding information and therefore functional groups within the sample. This provides the potential for understanding geographic, spatial and species-specific variation in plant biochemistry. Here we demonstrate the use of mid-FTIR microspectroscopy for the analysis of Drosera rotundifolia herbaria specimens, which were collected 100 years apart from different locations. Principal component and hierarchical clustering analysis enabled successful differentiation between three main regions on the plant (lamina, tentacle stalk and tentacle head), and between the different specimens. Lipids and protein spectral regions were particularly sensitive differentiators of plant tissues. Differences between the different sets of specimens were smaller. This study demonstrates that relevant information can be extracted from herbarium specimens using FTIR, with little impact on the specimens. FTIR therefore provides the potential as a powerful tool to unlock historic information within herbaria.

Sample Collection

Four D. rotundifolia herbarium specimens were used in this study (Table 1, Figure S1). Two of the specimens were plants collected in 2021 from the same location, at the same time by the same collector. The other two specimens were collected from a different location but were collected at the same place on the same date in 1934. All samples were in dried form with no observable moisture.

FTIR

Specimens were carefully removed from the herbarium sheet and placed onto the sample stage. FTIR spectra were taken for each of three tissue types (Fig. 1: lamina, tentacle stalk, tentacle head), on two leaves, for each of the four specimens. In each area, three replica FTIR spectra maps were produced. Each map consisted of roughly 15 points depending on area size (minimum = 8, maximum = 20). Infrared spectra were taken using a Thermo Nicolet iN10mx spectrometer fitted with a liquid-nitrogen-cooled MCT detector. Spectra were captured in transmission mode, with an 80 × 80 µm aperture size and 4 cm-1 resolution. Background spectra were taken in air before each sample, with 64 scans averaged at each map point; samples were suspended enabling direct transmission of IR.