P9: Isolation of a Citrobacter freundii mutant capable of elevated 3-hydroxypropionaldehyde production

The specialty chemical 3-hydroxypropionaldehyde is commercially valuable in the production of plastics. The isolation of a mutant strain of Citrobacter freundii ATCC 8090 capable of elevated 3-hydroxypropionaldehyde production from glycerol using buffered cells was sought. The mutant strain was isolated using ethylmethane sulfonate mutagenesis and a selection protocol. The selection protocol involved screening the mutagenized bacterial cells on solid minimal medium containing 5% glycerol. Colonies were picked onto duplicate solid minimal medium plates and one plate was stained with a solution of 1% phloroglucinol. Those colonies stained red by the phloroglucinol solution on the plates were picked for further screening. All possible mutants were grown in a complex medium containing 5% glycerol for 24 hours at 28^oC. The cells were collected by centrifugation and resuspended in a neutral phosphate buffer containing 5% glycerol and semicarbazide hydrochloride. The cell suspensions were shaken for 24 hours at 28^oC. After the cells were collected by centrifugation, the resultant supernatant of each suspension was spectrophotometrically assayed for 3-hydroxypropionaldehyde. One mutant strain was found to produce a several-fold higher 3-hydroxypropionaldehyde concentration than did ATCC 8090 when grown on 5% glycerol. When the mutant strain was grown on 10% glycerol, the aldehyde level produced by the mutant strain decreased but was still higher than the level produced by its parent strain. In conclusion, it was shown that a mutant of C. freundii could be isolated that was capable of elevated 3-hydroxypropionaldehyde from glycerol.