The present study reports the identification of two of the major unknown metabolites produced by P. scabrosum to be penigequinolone A and B when grown on Czapek Yeast Extract agar (CYA). Evaluation of the 400 MHz 1H and 100 MHz 13C NMR spectral data in CDCl3, including 2D experiments (DEPT, HMBC, NOESY), mass spectroscopy data (probe EIMS showing M` at m/z 467) and elemental microanalysis (C27H33O6 N) established what was considered a single compound, to be identical to the diastereomerical mixture of penigequinolone A and B described by Kimura et al. (1996). A total of 16 isolates of P. scabrosum were cultivated on CYA, and screened for secondary metabolite production using the plug extraction procedure and HPLC analysis. HPLC analysis demonstrated the metabolite profiles of the 16 isolates of P. scabrosum to be qualitatively very similar, each displaying penigequinolones A and B as major peaks together with cyclopenin, cyclopenol and viridicatin. The penigequinolones can be observed as ions at m/z 468 (M#H`) in the ES-MS analysis of the extracts, thus contributing to the unique mass profile of P. scabrosum when compared to other penicillia.