An adequate supply of standard reference material for paralytic shellfish toxins (PSTs) is critical for the accurate chemical quantification using high performance liquid chromatography (HPLC) with fluorescent detection, liquid chromatography-tandem mass spectrometry (LC-MS/MS), biological analysis of these toxins using enzyme-linked immunosorbent assay (ELISA), and immunochromatography. Large batch cultivation for the chain forming species G. catenatum, producers of PSTs of N-sulfocarbamoyl-11-hydroxysulfate toxins (C1 and C2), gonyautoxin 5 (GTX5) and gonyautoxin 6 (GTX6), was investigated using 10 L round-bottom flasks with aeration for the production of GTX5 and GTX6. Aeration rates of 200 mL/min and 500 mL/min were compared, demonstrating that the 500 mL/min aeration rate was adequate to eliminate aggregation of cells. The highest cell density of G. catenatum in 500 mL/min aeration treatment was 9,878 ± 2,617 cells/ml on day 28. Total toxin yield during 10 L cultivation with 500 mL/min aeration was calculated at 30.9 ± 3.6 µmol on day 25, with GTX5 and GTX6 calculated at 3.9 ± 0.7 µmol and 11.4 ± 1.4 µmol, respectively. This simple aeration method will contribute to the more efficient production of PST reference materials.