NPid: an Automatic Approach to Rapid Identification of Known Natural Products in the Crude Extract of Crabapple Based on 2D ¹H–¹³C Heteronuclear Correlation Spectra of the Extract Mixture

An automatic approach to identification of natural products (NPid) in complex extracts by exploring pure shift HSQC (psHSQC) and H2BC spectra of the mixture is developed, which integrated information on chemical shifts (CS), adjacent relationships (AR) and peak intensities (PI) of ¹H–¹³C groups for identification of candidate natural product in a customized NMR database. A weighted comprehensive score is calculated for each candidate from the values of CS, AR and PI to rate the likelihood of its existence in the complex mixture. Using the crude extract of crabapple (<i>Malus fusca</i>) as an example, a customized NMR database of natural products from plants of the genus <i>Malus</i> was constructed. The performance of NPid was first evaluated using simulated data in four scenarios, that is, for identification of structurally similar natural products, identification of natural products with part of peaks missing in psHSQC due to low concentration, without available adjacent relationship information, or without useful peak intensity information. The false positive and false negative rates of the natural products identified by NPid were estimated by Monte Carlo simulation. It shows that AR and PI can effectively reduce the false positive rate of identification. Proof of concept of the proposed method was elucidated on a model mixture consisting of 10 known natural products. Application of this method was then demonstrated on an authentic sample of crude extract of crabapple and 19 known natural products were successfully identified and confirmed by standard spiking.