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Background The young shoots of Rubus idaeus are traditionally used as a herbal remedy in common cold, fever and flu-like infections yet there is no research concerning this plant material. The aim of the study was to evaluate the chemical composition and biological properties of raspberry shoots from 11 cultivar varieties. Methods The methanol extracts were subjected to chromatographic analysis using HPLC-DAD-ESIMS, and two-dimensional 'comprehensive' LCxLC techniques. The biological activity of the shoot extract from the 'Willamette' cultivar variety was evaluated. Antioxidant activity was tested using DPPH and phosphomolybdenum assay. Antimicrobial activity was estimated towards 15 strains of human pathogenic bacteria using broth microdilution method. Cytotoxic activity was tested using MTT cell viability assay. Results The dominating compounds identified in the shoots of R. idaeus were ellagic acid (26.1 - 106.8 mg/100 g) and sanguiin H-6 (139.2 - 633.1 mg/100 g). The best separation of compounds present in the analysed polyphenol complex, was achieved by 'comprehensive' LCxLC method using Nucleodur Sphinx RP column in the first dimension and Chromolith Performance column in the second dimension. The shoot extract was found to be a strong antioxidant (EC50 19.4 μg/ml, AAE 427.94 mg/g) and displayed the strongest bactericidal properties towards Corynebacterium diphtheriae. The extract revealed higher cytotoxic activity towards the HL-60 cells (IC50 110 μg/ml) than HeLa (IC50 300 μg/ml). Conclusions The shoots of R. idaeus stand out as a valuable source of sanguiin H-6 and ellagic acid and possess a number of biological properties including antioxidative, antimicrobial and cytotoxic. [ABSTRACT FROM AUTHOR]