Qihe Chen

Optimization of medium composition for the production of elastase by Bacillus sp. EL31410 with response surface methodology

Abstract A newly isolated strain EL31410, which produced elastase (E.C3.4.4.7) with a high elastolytic activity was identified as Bacillus sp . The optimal conditions for the enzymatic reaction occurred at pH 7.4 and 37°C. Glucose and Casein are the best carbon and nitrogen sources for enzyme production. Addition of corn steep flour can affect the bacterium growth and elastase production. A fractional factorial design was applied to study the main factors that affect the enzyme production and the central composite experimental design and response surface methodology were adopted to derive a statistical model for the effect of casein and corn steep flour on elastase production. These experimental results showed that the corn steep flour had a positive effect and the high casein had a negative effect on enzyme production. An initial concentration of 1.15%(w/v) casein and 0.61%(w/v) corn steep flour were found to be best for the elastase production in batch culture. In addition, the time course of elastase production in the optimized medium composition was also described.

Optimization of ultrasonic-assisted extraction (UAE) of betulin from white birch bark using response surface methodology.

Betulin is an abundant naturally occurring triterpene, which makes it a potentially important raw material for a precursor of biologically active compounds. The objective of the current study was to determine the optimum UAE conditions for betulin from B. papyfera bark. The optimum conditions were evaluated with fractional factorial design and optimized using response surface methodology. High yields of betulin were observed from white birch bark by UAE technology. The solvent concentration and the ratio of material to solvent were the most significant parameters on betulin extraction as evaluated through FFD. The extraction conditions were further investigated with central composite design. The fitted second-order model revealed that the optimal conditions consisted of 98% ethonal concentration, 1:42 the ratio of white birch bark to solvent, extraction temperature 50 degrees C, ultrasonic frequency 5kHz and extraction time 3h. Under the optimized condition, the maximum productivity of betulin predicted is 23.17%. The extraction productivity and purity of betulin under the optimized extraction conditions were great higher than that of the non-optimized condition. The present study demonstrates that ultrasound is a great efficiency tool for the fast extraction of betulin from white birch bark.

Isolation and characterisation of an oxygen, acid and bile resistant Bifidobacterium animalis subsp. lactis Qq08

BACKGROUND Currently, bifidobacteria are recognised as one of the most important bacteria used as probiotics, which promote human health. However, their commercial application has been limited by their anaerobic nature. The purpose of this study was to select an oxygen, acid and bile resistant strain of bifidobacterium for use as a new probiotic. RESULTS A total of 10 strains of bifidobacteria from different sources were analysed for their relative bacterial growth ratio (RBGR) in different oxygen concentrations. Three strains with high RBGR values were selected and their survival rates in acid environment and bile salt conditions were investigated in vitro. One strain showed high tolerance to low pH, giving a survival rate of 84% at pH 2 after 4 h incubation, and high tolerance to bile, more than 90% after 4.5 h incubation at 0.01 g mL(-1) bile concentration. This strain was identified as Bifidobacterium animalis subsp. lactis strain Qq08 based on polyphasic taxonomy approaches, such as phenotype analysis and 16S rRNA and 16S to 23S internally transcribed spacer sequence analyses. CONCLUSION We isolated an aerotolerant bifidobacterium and identified it as Bifidobacterium animalis subsp. lactis Qq08. This strain has characteristics more favourable than the commercial probiotic strain, Bifidobacterium lactis Bb12.

Optimization of acidic extraction of astaxanthin from Phaffia rhodozyma

Optimization of a process for extracting astaxanthin from Phaffia rhodozyma by acidic method was investigated, regarding several extraction factors such as acids, organic solvents, temperature and time. Fractional factorial design, central composite design and response surface methodology were used to derive a statistically optimal model, which corresponded to the following optimal condition: concentration of lactic acid at 5.55 mol/L, ratio of ethanol to yeast dry weight at 20.25 ml/g, temperature for cell-disruption at 30 °C, and extraction time for 3 min. Under this condition, astaxanthin and the total carotenoids could be extracted in amounts of 1294.7 μg/g and 1516.0 μg/g, respectively. This acidic method has advantages such as high extraction efficiency, low chemical toxicity and no special requirement of instruments. Therefore, it might be a more feasible and practical method for industrial practice.

Studies on optimization of nitrogen sources for astaxanthin production by Phaffia rhodozyma

Fermentation of Phaffia rhodozyma is a major method for producing astaxanthin, an important pigment with industrial and pharmaceutical application. To improve astaxanthin productivity, single factor and mixture design experiments were used to investigate the effects of nitrogen source on Phaffia rhodozyma cultivation and astaxanthin production. Results of single factor experiments showed nitrogen source could significantly affect P. rhodozyma cultivation with respect to carbon source utilization, yeast growth and astaxanthin accumulation. Further studies of mixture design experiments using (NH4)2SO4, KNO3 and beef extract as nitrogen sources indicated that the proportion of three nitrogen sources was very important to astaxanthin production. Validation experiments showed that the optimal nitrogen source was composed of 0.28 g/L (NH4)2SO4, 0.49 g/L KNO3 and 1.19 g/L beef extract. The kinetic characteristics of batch cultivation were investigated in a 5-L pH-stat fermentor. The maximum amount of biomass and highest astaxanthin yield in terms of volume and in terms of biomass were 7.71 mg/L and 1.00 mg/g, respectively.

Biotransformation optimization of betulin into betulinic acid production catalysed by cultured Armillaria luteo-virens Sacc ZJUQH100-6 cells

Aims:  Betulinic acid has attracted attention in terms of its important biological and pharmacological characteristics. The main objective of this work was to optimize the variables of biotransformation process in order to enhance betulinic acid production from betulin catalysed by fungus Armillaria luteo‐virens Sacc ZJUQH100‐6.

Optimization of biotransformation from phytosterol to androstenedione by a mutant Mycobacterium neoaurum ZJUVN-08

Biotransformation of phytosterol (PS) by a newly isolated mutant Mycobacterium neoaurum ZJUVN-08 to produce androstenedione has been investigated in this paper. The parameters of the biotransformation process were optimized using fractional factorial design and response surface methodology. Androstenedione was the sole product in the fermentation broth catalyzed by the mutant M. neoaurum ZJUVN-08 strain. Results showed that molar ratio of hydroxypropyl-β-cyclodextrin (HP-β-CD) to PS and substrate concentrations were the two most significant factors affecting androstenedione production. By analyzing the statistical model of three-dimensional surface plot, the optimal process conditions were observed at 0.1 g/L inducer, pH 7.0, molar ratio of HP-β-CD to PS 1.92:1, 8.98 g/L PS, and at 120 h of incubation time. Under these conditions, the maximum androstenedione yield was 5.96 g/L and nearly the same with the non-optimized (5.99 g/L), while the maximum PS conversion rate was 94.69% which increased by 10.66% compared with the non-optimized (84.03%). The predicted optimum conditions from the mathematical model were in agreement with the verification experimental results. It is considered that response surface methodology was a powerful and efficient method to optimize the parameters of PS biotransformation process.

Hydrolyzates of silkworm pupae (Bombyx mori) protein is a new source of angiotensin I-converting enzyme inhibitory peptides (ACEIP).

Silkworm pupae protein is a good source of high quality protein. The hydrolyzates of silkworm pupae protein catalyzed by neutrase, pepsin, acidic protease (Asperqiius usamii NO. 537), flavourzyme, alcalase, and trypsin with inhibitory activity on angiotensin I-converting enzyme (ACE) were identified by HPLC. The hydrolyzates catalyzed by acidic protease exerted the highest inhibitory activity on ACE. The hydrolyzing conditions were optimized by one-factor, factional factorial (FFD), and center composite (CCD) design methods, and response surface methodology (RSM). Statistical analyses showed that regression of the second-order model equation is suitable to describe ACE inhibitory bioactivity. The predicted inhibitory activity of hydrolyzates on ACE was 73.5 % at a concentration of 2.0 mg/ml. Optimized RSM technique decreased IC(50) of hydrolyzates inhibiting ACE to 1.4 mg/mL from 2.5 mg/ml. The molecular weight of the components of the hydrolyzates with inhibitory activity on ACE varied from less than 500 to about 1000 Da by ultra-filter analysis. These studies suggest that hydrolyzates of silkworm protein contain ACE inhibitory activity that could form a potential source of ACE inhibitor drugs.

Preparative isolation and purification of xanthohumol from hops (Humulus lupulus L.) by high-speed counter-current chromatography

Xanthohumol (XN) and related prenylflavonoids are the main bioactive components of hops (Humulus lupulus L.). The current work is to investigate the use of high-speed counter-current chromatography (HSCCC) in search for high isolation of xanthohumol from hops. A solvent system consisted of n-hexane-ethyl acetate-methanol-water at a volume ratio of 5:5:4:3 was employed. The results demonstrated that the constructed method could be well applied for the isolation of xanthohumol from hops extract. After HSCCC isolation procedure, the purity of xanthohumol was over 95% assayed by HPLC and the yield of extraction was 93.60%. The chemical structure identification of xanthohumol was carried out by UV, (1)H NMR and (13)C NMR. The present results demonstrated that xanthohumol could be efficiently obtained using a single HSCCC step from H. lupulus L. extract.

Optimization of culture medium compositions for gellan gum production by a halobacterium Sphingomonas paucimobilis.

The effect of culture medium compositions on gellan gum production produced by fermentation with a halobacterium Sphingomonas paucimobilis QHZJUJW CGMCC2428 was studied. In this work, a fractional factorial design was applied to investigate the main factors that affected gellan gum production by S. paucimobilis QHZJUJW CGMCC2428. Sucrose was the best carbon source for gellan gum and peptone displayed better inducing effect. Central composite design and response surface methodology were adopted to derive a statistical model for optimizing submerged culture medium composition. These experimental results showed that the optimum culture medium for producing gellan gum was composed of 40.00 (w/v) sucrose, 3.00% peptone (w/v), MgSO4 (w/v), 9.20% KH2PO4 (w/v), 7.50% Na2HPO4 (w/v), 4.30% K2SO4 (w/v), pH 6.8-7.0. The maximal gellan gum was 19.89±0.68 g/L, which was agreed closely with the predicated value (20.12 g/L). After incubated for 72 h under the optimized culture medium in 5-L bioreactor, the gellan gum fermentation reached about 19.90±0.68 g/L, which was higher than that in the initial cultivation medium.