Supercritical CO 2 was used to fractionate a mixture of monoacylglycerols (MAGs), diacylgylcerols (DAGs) and triacylglycerols (TAGs) using an eight-foot packed column which was kept under thermal gradient conditions and operated semi-continuously. Fractionation of the glyceride mixtures was affected by both their solubility in CO 2 and respective vapor pressure enhancement. The feed material used, approximating a feed stream used in the industrial enrichment of MAGs, was composed of 48.7 wt.% MAG, 46.3 wt.% DAG and 5 wt.% TAG, having main acyl chain compositions of 6 wt.% C16:0, 23 wt.% C18:0 and 59 wt.% C18:1. The resultant top product from the fractionation column had MAG concentrations as high as 90 wt.% and no TAG, which is comparable to that produced by molecular distillation. The effects of pressure, temperature gradient, CO 2 flow rate and feed composition on the separation efficiency and yields were investigated. Increasing the column pressure from 172 to 344 bar or the density of CO 2 from 420 to 720 kg m −3 increased the yield of the top product. However, with those increases, the supercritical fluid became less selective, with the MAG concentration decreasing from 84 to 56 wt.%. At 207 bar, a linear temperature gradient varying from 65 to 95 °C gave the best selectivity when compared to other temperature gradient schemes. Increasing the CO 2 flux from 1.8 to 10.3 g min −1 cm −2 resulted in little change in the MAG concentration, but the top-product yield decreased from 14.4 to 2.3 g kg −1 CO 2. As the MAG concentration in the feed mixture increased from 48.7 to 63 wt.%, the MAG concentration in the top product (at 207 bar and 65–95 °C) increased from 79.4 to 86 wt.%.