The combretazet beta-lactams are cis-restricted analogues of combretastatin A-4, chemically manipulated to enhance their in vitro and in vivo stability. They are extremely potent anti-proliferative agents targeting the colchicine-binding site of tubulin, as microtubule destabilisers leading to G₂/M phase arrest, karyorrhexis and ultimately, apoptosis and cell death. 3-Hydroxyl combretazets demonstrate excellent IC₅₀ values in subnanomolar range across a panel of cancer cell lines. We have previously reported chiral diastereomeric resolution of b-lactam racemates using chiral derivatising reagent N-Boc-L-proline to afford enantiopure b-lactams, albeit with large process mass intensities (PMI) of approximately 150,000 kg/kg and modest yields (5-10%), insufficient for progression toward in vivo pre-clinical toxicology studies. The 3S,4S enantiomer was isolated in >90% enantiomeric excess (ee). However, poor diastereomeric excess was obtained for the second eluting 3R,4R diastereomer yielding equally poor ee (50-60%) for its corresponding enantiomer. The process was labour and solvent intensive. Committed to the design of a sustainable, green and safer resolution process, biocatalysis using lipase chemoenzymatic mediated kinetic resolution (KR) toward combretazet enantiomers was investigated.Candida antarctica lipase B (CAL-B) catalysed KR of 3-acetoxy beta-lactams toward their corresponding enantioenriched 3-hydroxyl 3S,4S beta-lactams in 70-80% ee, comparable to values achieved via diastereomeric resolution. PMI was reduced by 1,300-30,000 fold compared to diastereomeric resolution. Candida antarctica KR of combretazet b-lactams is an accessible enantioseparation strategy, reducing environmental impact while overcoming issues associated with poor yield, with KR yielding sufficient quantities of beta-lactam enantiomers for pre-clinical in vivo toxicological evaluation.