Two major anthocyanins, cyanidin 3-O-glucoside and 3-O-rutinoside, were isolated from the black flowers of Cosmos atrosanguineus cultivar 'Choco Mocha', together with three minor anthocyanins, cyanidin 3-O-malonylglucoside, pelargonidin 3-O-glucoside and 3-O-rutinoside. A chalcone, butein 4'-O-glucoside and three minor flavanones were isolated from the red flowers of C. atrosanguineis x C. sulphureus cultivar 'Rouge Rouge'. The anthocyanins and chalcone accumulation of cultivar 'Choco Mocha' and its hybrid cultivars 'Brown Rouge', 'Forte Rouge', 'Rouge Rouge' and 'Noel Rouge' was surveyed by quantitative HPLC. Total anthocyanins of black flower cultivars 'Choco Mocha' and 'Brown Rouge' were 3-4-folds higher than that of the red flower cultivar 'Noel Rouge'. On the other hand, total chalcone of 'Noel Rouge' was 10-77-folds higher compared with those of other cultivars, 'Brown Rouge', 'Forte Rouge' and 'Rouge Rouge'. It was shown that the flower color variations from red to black of Chocolate Cosmos and its hybrids are due to the difference in the relative amounts of anthocyanins and chalcone.