Chemotaxonomy is a method of classifying plants based on their chemical features. This can be used in combination with genomics for robust species identification. The objective of the current study was to test the suitability of preserved specimens for species delimitation using total metabolomic profiles. Herbarium voucher specimen leaf samples from three species of Banksia L. f. were used for the current experiment. This included Banksia neoanglica (A.S.George) Stimpson & J.J.Bruhl, Banksia penicillata (A.S.George) K.R.Thiele and Banksia robur Cav. Dry leaf samples were homogenized and the methanolic extracts were analyzed using liquid chromatography-tandem mass spectrometry and the resulting mass spectral data were analyzed using multi-variate statistical analysis. To test the suitability of herbarium dry specimens for species delimitation, we conducted hierarchical cluster analysis based on their total metabolomic profiles. Based on the hierarchical cluster analysis results there were three distinct clusters for B. neoanglica, B. robur and B. penicillata. This was further verified by analysis of similarity values between the chemical profiles of three species ( R= 0.925, p < 0.05). It should also be noted that the voucher specimens from each species that were used in this analysis were collected from different localities and preserved at different time scales. Our results therefore show that regardless of preservation age and collection locality, herbarium specimens are chemically informative for species delimitation.