tSemi-preparative HPLC is one of the main techniques used for the purification of natural products (NPs).Generally, the sample has to be solubilized in organic solvent and injected on column through a loop valve.Since the solubility of crude natural extracts is often limited, a high solvent volume is needed for injection.This significantly compromises the resolution and increases the risk of overpressure in the system. Toovercome this problem, a dry load injection procedure was developed to ensure optimum resolution evenat high sample loading. The approach was first validated with a representative mixture of NPs standards,and successfully applied to two representative crude plant extracts: the dichloromethane extract ofAnnacardium occidentale and the methanolic extract of Hypericum perforatum. In all cases, the dry loadinginjection setup enabled an efficient introduction of the samples in the semi-preparative HPLC system.Different overload conditions of the columns were tested and the results demonstrated the robustness ofthe method and the possibility of applying it with a limited loss of resolution compared to liquid injectionand without increasing pressure. The chromatographic resolutions were close to those obtained at theanalytical level and separation were of much better quality when compared to liquid injection. Thisapproach is especially relevant when purifying compounds isolated with high resolution from extractsthat are poorly soluble in low volume of injection solvent due to the presence of lipophilic compoundsand are thus not compatible for loop injection in typical reversed phase conditions. In addition, thedry load setup was also found to be useful when relatively polar components have to be separated inreversed phase conditions. In this case, loop injection with methanol generates strong peak distortionand broadening, while the dry load injection affords symmetrical peaks.