The method has been proposed for the spectrophotometric determination of antibiotics of the cephalosporin (cefuroxime, ceftriaxone, cefotaxime, cefazolin) and penicillin (amoxicillin) groups using chemometric data processing (method of partial least squares, PLS). The absorption spectra of 24 binary mixtures of “cephalosporin – cephalosporin” and 16 binary mixtures of “cephalosporin – amoxicillin” with various concentrations in the range of 3–27 and 5–140 μg/ml have been recorded. The optimal number of latent variables for each chemometric method (PLS-1 and PLS-2) has been selected, which ranged from 2 to 7 depending on the type of model and the nature of the antibiotics. The smallest root mean square calibration errors (RMSEC) have been found for the ceftriaxone–cefazolin system, which have been 0.09 and 0.08 for PLS-1 and PLS-2, respectively. The smallest root mean square errors of prediction (RMSEP) for the mixture “cefuroxime – amoxicillin” have been 0.07 for both methods (PLS-1 and PLS-2). It has been established that equations of linear relationships “measured-predicted” are characterized by a regression coeffi cient and the square of the approximation coeffi cient close to 1; which indicates the high quality of the models. The relative error of determination in test mixtures has been 0.4–6.2% by the PLS-1 method for “cephalosporin – cephalosporin” mixtures, and 0.07–6.0% by the PLS-2 method. At the same time, there is no signifi cant diff erence in the error in determining the components of the mixture, in contrast to the “cefuroxime – amoxicillin” system, for which the error is 0.1–1.1% for the fi rst component and 2.6–14.7% for the second component using the PLS-1 method and 0.2–1.2% for the fi rst component and 4.3–13.3% for the second component using the PLS-2 method. Thus, it has been shown that the use of chemometric methods ensures high accuracy of the spectrophotometric determination of the studied antibiotics with overlapping absorption bands in their mixtures.