TY - JOUR
T1 - Fed-batch bioconversion of indene to cis-inandiol
AU - Amanullah, A
AU - Hewitt, Christopher
AU - Nienow, Alvin
AU - Lee, C
AU - Chartrain, M
AU - Buckland, BC
AU - Drew, SW
AU - Woodley, JM
PY - 2002/12/2
Y1 - 2002/12/2
N2 - The bioconversion of indene to cis-(1S,2R) indandiol, a key intermediate in the synthesis of Merck's HIV protease inhibitor, CRIXIVAN(TM) can be achieved during the growth of a Rhodococcus strain. In a previous study, we reported on the application of multi-parameter flow cytometry for the measurement of indene toxicity to the strain, and found that concentrations up to 0.25 g/l of indene (0.037 g indene/g dry cell weight) in batch bioconversions did not influence cell physiology. Using this information, this study reports on the implementation of a single phase indene fed-batch bioconversion. Cytoplasmic membrane (membrane) integrity and membrane polarisation of a large number of cells were measured during such bioconversions using multi-parameter flow cytometry and compared to a control in order to assess any toxic effects of indene feeding. The results indicate that indene supply at a rate of 0.1 g/l/h is feasible without any deleterious effects on cell physiology. The delay in indene metabolism was significantly shorter, with lower concentrations of by-product formation, when it was added to the culture in the stationary phase than when it was added at the beginning of the exponential phase of the fermentation. cis-Indandiol production rates could be enhanced from 20 mg/l/h, in a previously reported silicone oil two-liquid phase system, up to 200 mg/l/h by a combination of suitable indene feeding rates in the stationary phase and operating with a high biomass concentration to limit the effects of toxicity. In addition, the yield of cis-indandiol on indene (g/g) was higher at 0.48 in the single phase system compared to 0.20 in the two-liquid phase system. However, the final concentration of cis-indandiol was considerably lower, possibly as a result of higher dehydrogenase activity resulting in an increased transformation of cis-indandiol to 1-keto-2-hydroxy indan. This study has demonstrated that it is feasible to feed indene directly in the stationary phase of the bioconversion using high biomass concentrations to obtain enhanced cis-indandiol formation rates as well as yields based on indene utilisation compared to a two-phase silicone oil system. (C) 2002 Elsevier Science Inc. All rights reserved.
AB - The bioconversion of indene to cis-(1S,2R) indandiol, a key intermediate in the synthesis of Merck's HIV protease inhibitor, CRIXIVAN(TM) can be achieved during the growth of a Rhodococcus strain. In a previous study, we reported on the application of multi-parameter flow cytometry for the measurement of indene toxicity to the strain, and found that concentrations up to 0.25 g/l of indene (0.037 g indene/g dry cell weight) in batch bioconversions did not influence cell physiology. Using this information, this study reports on the implementation of a single phase indene fed-batch bioconversion. Cytoplasmic membrane (membrane) integrity and membrane polarisation of a large number of cells were measured during such bioconversions using multi-parameter flow cytometry and compared to a control in order to assess any toxic effects of indene feeding. The results indicate that indene supply at a rate of 0.1 g/l/h is feasible without any deleterious effects on cell physiology. The delay in indene metabolism was significantly shorter, with lower concentrations of by-product formation, when it was added to the culture in the stationary phase than when it was added at the beginning of the exponential phase of the fermentation. cis-Indandiol production rates could be enhanced from 20 mg/l/h, in a previously reported silicone oil two-liquid phase system, up to 200 mg/l/h by a combination of suitable indene feeding rates in the stationary phase and operating with a high biomass concentration to limit the effects of toxicity. In addition, the yield of cis-indandiol on indene (g/g) was higher at 0.48 in the single phase system compared to 0.20 in the two-liquid phase system. However, the final concentration of cis-indandiol was considerably lower, possibly as a result of higher dehydrogenase activity resulting in an increased transformation of cis-indandiol to 1-keto-2-hydroxy indan. This study has demonstrated that it is feasible to feed indene directly in the stationary phase of the bioconversion using high biomass concentrations to obtain enhanced cis-indandiol formation rates as well as yields based on indene utilisation compared to a two-phase silicone oil system. (C) 2002 Elsevier Science Inc. All rights reserved.
KW - cis-indandiol
KW - Rhodococcus
KW - HIV protease inhibitor
KW - indene
KW - fed-batch bioconversion
KW - flow cytometry
UR - http://www.scopus.com/inward/record.url?scp=2242452373&partnerID=8YFLogxK
U2 - 10.1016/S0141-0229(02)00183-7
DO - 10.1016/S0141-0229(02)00183-7
M3 - Article
VL - 31
SP - 954
EP - 967
JO - Enzyme and Microbial Technology
JF - Enzyme and Microbial Technology
IS - 7
ER -