Biomedical subjects

Shu-Wei Chang

Publications and source records attributed to Shu-Wei Chang.

7 recordsLinked to original sources

PubMed2006

Optimal formation of hexyl laurate by Lipozyme IM-77 in solvent-free system.

A medium-chain ester, hexyl laurate, with fruity flavor is primarily used in personal care formulations as an important emollient for cosmetic applications. To conform to the "natural" interests of consumers, the ability of immobilized lipase from Rhizomucor miehei (Lipozyme IM-77) to catalyze the direct esterification of hexanol and lauric acid by using a solvent-free system was investigated in this study. Response surface methodology (RSM) and four-factor-five-level central composite rotatable design (CCRD) were employed to evaluate the effects of synthesis parameters, such as reaction time (10-50 min), temperature (45-85 degrees C), lipase amount (10-30 mg/volume; 0.077-0.231 batch acidolysis units of Novo (BAUN), and pH memory (5-9), on percentage molar conversion of hexyl laurate by lipase-catalyzed direct esterification. Reaction time, temperature, and enzyme amount had significant effects on percent molar conversion. On the basis of ridge maximum analysis, the optimum synthesis conditions for hexyl laurate were a reaction time of 40.6 min, a temperature of 58.2 degrees C, an enzyme amount of 25.4 mg/volume (0.196 BAUN), and a pH memory of 5.9. The predicted percentage molar conversion of hexyl laurate was 69.7 +/- 1.4%.

Enzymes, Immobilized↗

PubMed2006

Efficient production of active recombinant Candida rugosa LIP3 lipase in Pichia pastoris and biochemical characterization of the purified enzyme.

Candida rugosa lipase (CRL), an important industrial enzyme, possesses several different isoforms encoded by the high-identity lip gene family (lip1 to lip7). In this study, an additional N-terminal peptide in front of the lip3 gene was removed by PCR, and the 18 nonuniversal serine codons (CTG) of the lip3 gene were converted into universal serine codons (TCT) by means of an overlap extension PCR-based multiple-site-directed mutagenesis to express an active recombinant LIP3 in the yeast Pichia pastoris. The regional synthetic DNA fragment (339 bp) is first recombined by primer assembly with 20 overlapping nucleotides, followed by specific overlap extension PCR with outside primers containing restriction enzyme sites for directional cloning into the pGAPZalphaC vector. The results show that the production yield (0.687 unit/mL) of N-fused lip3 (nflip3) has an overall improvement of 69-fold relative to that (0.01 unit/mL) of lip3 and of 52-fold (0.47 unit/mL) of codon-optimized lip3 (colip3) relative to that (0.01 unit/mL) of non-codon-optimized lip3 (lip3), with the cultivation time set at 5 days. This finding demonstrates that the reservation of the N terminus and the regional codon optimization of the lip3 gene fragment at the 5' end can greatly increase the expression level of recombinant LIP3 in the P. pastoris system. The purified recombinant LIP3 shows distinct biochemical properties compared with other isoforms.

Amino Acid Sequence↗

PubMed2006

Candida rugosa lipase LIP1-catalyzed transesterification to produce human milk fat substitute.

Structured lipids (SLs) containing palmitic and oleic acids were synthesized by transesterification of tripalmitin with either oleic acid or methyl oleate as acyl donor. This SL with palmitic acid at the sn-2 position and oleic acid at sn-1,3 positions is similar in structure to human milk fat triacylglycerol. LIP1, an isoform of Candida rugosa lipase (CRL), was used as biocatalyst. The effects of reaction temperature, substrate molar ratio, and time on incorporation of oleic acid were investigated. Reaction time and temperature were set at 6, 12, and 24 h, and 35, 45, and 55 degrees C, respectively. Substrate molar ratio was varied from 1:1 to 1:4. The highest incorporation of oleic acid (37.7%) was at 45 degrees C with methyl oleate as acyl donor. Oleic acid resulted in slightly lesser (26.3%) incorporation. Generally, higher percentage incorporation of oleic acid was observed with methyl oleate (transesterification) than with oleic acid (acidolysis). In both cases percentage incorporation increased with reaction time. Incorporation decreased with increase in temperature above 45 degrees C. Initially, oleic acid incorporation increased with increase in substrate molar ratio up to 1:3. LIP1 was also compared with Lipozyme RM IM as biocatalysts. The tested reaction parameters were selected on the basis of maximum incorporation of C18:1 obtained during optimization of LIP1 reaction conditions. Reaction temperature was maintained at 45, 55, and 65 degrees C. Lipozyme RM IM gave highest oleic acid incorporation (49.4%) at 65 degrees C with methyl oleate as acyl donor. Statistically significant (P < 0.05) differences were observed for both enzymes. SL prepared using Lipozyme RM IM may be more suitable for possible use in human milk fat substitutes.

Animals↗

PubMed2006

Codon optimization of Candida rugosa lip1 gene for improving expression in Pichia pastoris and biochemical characterization of the purified recombinant LIP1 lipase.

An important industrial enzyme, Candida rugosa lipase (CRL) possesses several different isoforms encoded by the lip gene family (lip1-lip7), in which the recombinant LIP1 is the major form of the CRL multigene family. Previously, 19 of the nonuniversal serine codons (CTG) of the lip1 gene hav been successfully converted into universal serine codons (TCT) by overlap extension PCR-based multiple-site-directed mutagenesis to express an active recombinant LIP1 in the yeast Pichia pastoris. To improve the expression efficiency of recombinant LIP1 in P. pastoris, a regional synthetic gene fragment of lip1 near the 5' end of a transcript has been constructed to match P. pastoris-preferred codon usage for simple scale-up fermentation. The present results show that the production level (152 mg/L) of coLIP1 (codon-optimized LIP1) has an overall improvement of 4.6-fold relative to that (33 mg/L) of non-codon-optimized LIP1 with only half the cultivation time of P. pastoris. This finding demonstrates that the regional codon optimization the lip1 gene fragment at the 5' end can greatly increase the expression level of recombinant LIP1 in the P. pastoris system. More distinct biochemical properties of the purified recombinant LIP1 for further industrial applications are also determined and discussed in detail.

Base Sequence↗

PubMed2006

Optimized growth kinetics of Pichia pastoris and recombinant Candida rugosa LIP1 production by RSM.

A predictive model for Pichia pastoris expression of highly active recombinant Candida rugosa LIP1 was developed by combining the Gompertz function and response surface methodology (RSM) to evaluate the effect of yeast extract concentration, glucose concentration, temperature, and pH on specific responses. Each of the responses (maximum population densities, specific growth rate (mumax), protein concentration, and minimum lag phase duration) was determined using the modified Gompertz function. RSM and 4-factor-5-level central composite rotatable design (CCRD) were adopted to evaluate the effects of growth parameters, such as temperature (21.6-38.4 degrees C), glucose concentration (0.3-3.7%), yeast extract (0.16-1.84%), and pH (5.3-8.7) on the responses of P. pastoris growth kinetics. Based on ridge maximum analysis, the optimum population density conditions were: temperature 24.4 degrees C, glucose concentration 2.0%, yeast extract 1.5%, and pH 7.6. The optimum specific growth rate conditions were: temperature 28.9 degrees C, glucose concentration 2.0%, yeast extract 1.1%, and pH 6.9. The optimum protein concentration conditions were: temperature 24.2 degrees C, glucose concentration 1.9%, yeast extract 1.5%, and pH 7.6. Based on ridge minimum analysis, the minimal lag phase conditions were: temperature 32.3 degrees C, glucose concentration 2.1%, yeast extract 1.1%, and pH 5.4. For the predicted value, the maximum population density, specific growth rate, protein concentration, and minimum lag phase duration were 15.7 mg/ml, 3.4 h(-1), 0.78 mg/ml, and 4.2 h, and the actual values were 14.3 +/- 3.5 mg/ml, 3.6 +/- 0.6 h(-1), 0.72 +/- 0.2 mg/ml, and 4.4 +/- 1.6 h, respectively.

Candida↗

PubMed2004

Slow light in semiconductor quantum wells.

We demonstrate slow light via population oscillation in semiconductor quantum-well structures for the first time. A group velocity as low as 9600 m/s is inferred from the experimentally measured dispersive characteristics. The transparency window exhibits a bandwidth as large as 2 GHz.

Journal Article↗

PubMed2003

Application of solvent engineering to optimize lipase-catalyzed 1,3-diglyacylcerols by mixture response surface methodology.

1,3-Diacylglycerol (DAG) has been introduced in Japan as a cooking oil under the trade name of Econa to reduce body fat accumulation. Solvent engineering was applied to determine the optimum solvent mixtures for the lipase-catalyzed synthesis of 1,3-DAG by mixture response surface methodology. n-Hexane was required to maintain the lipase activity and the product selectivity could be adjusted by changing the hydrophobicity of reaction medium. The optimum yield (approximately 40%) of 1,3-DAG synthesis was obtained with n-hexane/octane (1:1, v/v).

Catalysis↗