The aerial parts and seeds of Sophora alopecuroides L. var. alopecuroides growing in Turkey were investigated for their alkaloid compositions and antimicrobial activities. The alkaloid extracts were analysed by capillary gas chromatography-mass spectrometry (GC-MS). The main components were identified as sophoridine (33.07%), matrine (24.03%), and sophocarpine (8.01%) in the aerial parts, and matrine (36.67%) and sophoridine (34.53%) in the seeds of the plant. The alkaloid extracts were also evaluated for their in vitro antibacterial and antifungal activities using microdilution techniques. The alkaloid extract of the aerial parts of S. alopecuroides var. alopecuroides presented significant activity against Staphylococcus aureus and Bacillus subtilis while the alkaloid extract of the seeds displayed good activity against Pseudomonas aeruginosa and Bacillus subtilis with minimum inhibitory concentrations (MICs) of 62.5 mu g/mL. In the antifungal assay, the alkaloid extract of the seeds of the plant showed significant activity against Candida krusei (MIC = 62.5 mu g/mL). To the best of our knowledge, the current work is the first to report the alkaloid profile and antimicrobial activity of S. alopecuroides var. alopecuroides growing in Turkey.