Separation of Nattokinase from Bacillus subtilis Fermentation Broth by Expanded Bed Adsorption with Mixed-mode Adsorbent

Lu Miao-Hua,Lin Dong-Qiang,Wu Yuan-Chun,Yun Jun-Xian,Mei Le-He,Yao Shan-Jing The Korean Society for Biotechnology and Bioengine 2005 Biotechnology and Bioprocess Engineering Vol.10 No.2

Mixed-mode hydrophobic/ionic matrices exhibit a salt-tolerant property for adsorbing target protein from high-ionic strength feedstock, which allows the application of undiluted feedstock via an expanded bed process. In the present work, a new type of mixed-mode adsorbent designed for expanded bed adsorption, Fastline $PRO^{\circledR}$, was challenged for the capture of nattokinase from the high ionic fermentation broth of Bacillus subtilis. Two important factors, pH and ion concentration, were investigated with regard to the performance of nattokinase ad-sorption. Under initial fermentation broth conditions (pH 6.6 and conductivity of 10 mS/cm) the adsorption capacity of nattokinase with Fastline PRO was high, with a maximum capacity of 5,350 U/mL adsorbent. The elution behaviors were investigated using packed bed adsorption experiments, which demonstrated that the effective desorption of nattokinase could be achieved by effecting a pH of 9.5. The biomass pulse response experiments were carried out in order to evaluate the biomass/adsorbent interactions between Bacillus subtilis cells and Fastline PRO, and to demonstrate a stable expanded bed in the feedstock containing Bacillus subtilis cells. Finally, an EBA process, utilizing mixed-mode Fastline PRO adsorbent, was optimized to capture nattokinase directly from the fermentation broth. The purification factor reached 12.3, thereby demonstrating the advantages of the mixed-mode EBA in enzyme separation.

Separation of Nattokinase from Bacillus subtilis Fermentation Broth by Expanded Bed Adsorption with Mixed-mode Adsorbent

Shan-Jing Yao,Miao-Hua Lu,Dong-Qiang Lin,Yuan-Chun Wu,Jun-Xian Yun,Le-He Mei 한국생물공학회 2005 Biotechnology and Bioprocess Engineering Vol.10 No.2

Mixed-mode hydrophobic/ionic matrices exhibit a salt-tolerant property for adsorbing target protein from high-ionic strength feedstock, which allows the application of undiluted feedstock via an expanded bed process. In the present work, a new type of mixed-mode adsorbent designed for expanded bed adsorption, Fastline PRO, was challenged for the capture of nattokinase from the high ionic fermentation broth of Bacillus subtilis. Two important factors, pH and ion concentration, were investigated with regard to the performance of nattokinase adsorption. Under initial fermentation broth conditions (pH 6.6 and conductivity of 10 mS/cm) the adsorption capacity of nattokinase with Fastline PRO was high, with a maximum capacity of 5,350 U/mL adsorbent. The elution behaviors were investigated using packed bed adsorption experiments, which demonstrated that the effective desorption of nattokinase could be achieved by effecting a pH of 9.5. The biomass pulse response experiments were carried out in order to evaluate the biomass/adsorbent interactions between Bacillus subtilis cells and Fastline PRO, and to demonstrate a stable expanded bed in the feedstock containing Bacillus subtilis cells. Finally, an EBA process, utilizing mixed-mode Fastline PRO adsorbent, was optimized to capture nattokinase directly from the fermentation broth. The purification factor reached 12.3, thereby demonstrating the advantages of the mixed-mode EBA in enzyme separation.

Emodin-Provoked Oxidative Stress Induces Apoptosis in Human Colon Cancer HCT116 Cells through a p53-Mitochondrial Apoptotic Pathway

Xie, Mei-Juan,Ma, Yi-Hua,Miao, Lin,Wang, Yan,Wang, Hai-Zhen,Xing, Ying-Ying,Xi, Tao,Lu, Yuan-Yuan Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.13

Emodin, a natural anthraquinone isolated from the traditional Chinese medicine Radix rhizoma Rhei, can induce apoptosis in many kinds of cancer cells. This study demonstrated that emodin induces apoptosis in human colon cancer HCT116 cells by provoking oxidative stress, which subsequently triggers a p53-mitochondrial apoptotic pathway. Emodin induced mitochondrial transmembrane potential loss, increase in Bax and decrease in Bcl-2 expression and mitochondrial translocation and release of cytochrome c to cytosol in HCT116 cells. In response to emodin-treatment, ROS increased rapidly, and subsequently p53 was overexpressed. Pretreatment with the antioxidant NAC diminished apoptosis and p53 overexpression induced by emodin. Transfecting p53 siRNA also attenuated apoptosis induced by emodin, Bax expression and mitochondrial translocation being reduced compared to treatment with emodin alone. Taken together, these results indicate that ROS is a trigger of emodin-induced apoptosis in HCT116 cells, and p53 expression increases under oxidative stress, leading to Bax-mediated mitochondrial apoptosis.

Genetic variations in the bitter taste receptor gene TAS2R38 are related to cigarette smoking behavior in Han Chinese smokers

Qi Fei-Yan,Zhu Zhou-Hai,Li Meng,Guan Ying,Peng Qi-Yuan,Lu She-Ming,Liu Zhi-Hua,Wang Ming-Feng,Miao Ming-Ming,Chen Zhang-Yu,Li Xue-Mei,Bai Jie,Yao Jian-Hua 한국유전학회 2022 Genes & Genomics Vol.44 No.11

Background: Smoking behavior is influenced by multiple genes, including the bitter taste gene TAS2R38. It has been reported that the correlation between TAS2R38 and smoking behavior has ethnicity-based differences. However, the TAS2R38 status in Chinese smokers is still unclear. Objective: This study aims to investigate the possible relationship between genetic variations in TAS2R38 (A49P, V262A and I296V) and smoking behaviors in the Han Chinese population. Methods: The haplotype analyses were performed and smoking behavior questionnaire was completed by 1271 individuals. Genetic association analyses for smoking behavior were analyzed using chi-square test. Further, for investigating the molecular mechanism of TAS2R38 variants effect on smoking behavior, we conducted TAS2R38-PAV and TAS2R38-AVI expression plasmids and tested the cellular calcium assay by cigarette smoke compounds stimulus in HEK293. Results: Significant associations of genetic variants within TAS2R38 were identified with smoking behavior. We found a higher PAV/PAV frequency than AVI/AVI in moderate and high nicotine dependence (FTND ≥ 4; X2 = 4.611, 1 df, p = 0.032) and strong cigarette smoke flavor intensity preference (X2 = 4.5383, 1 df, p = 0.033) in participants. Furthermore, in the in vitro cellular calcium assay, total particle matter (TPM), N-formylnornicotine and cotinine, existing in cigarette smoke, activated TAS2R38-PAV but not TAS2R38-AVI-transfected cells. Conclusion: Our data highlights that genetic variations in TAS2R38 are related to smoking behavior, especially nicotine dependence and cigarette smoke flavor intensity preference. Our findings may encourage further consideration of the taste process to identify individuals susceptible to nicotine dependence, particularly Han Chinese smokers.

Radiosensitivity Enhancement by Arsenic Trioxide in Conjunction with Hyperthermia in the EC-1 Esophageal Carcinoma Cell Line

Cui, Yan-Hui,Liang, Hai-Jun,Zhang, Qing-Qin,Li, Si-Qing,Li, Xiao-Rui,Huo, Xiao-Qing,Yang, Qing-Hui,Li, Wei-Wei,Gu, Jian-Fa,Hua, Qin-Liang,Lu, Ping,Miao, Zhan-Hui Asian Pacific Journal of Cancer Prevention 2012 Asian Pacific journal of cancer prevention Vol.13 No.4

Objective: To explore the effect on radiosensitivity of arsenic trioxide ($As_20_3$) in conjunction with hyperthermia on the esophageal carcinoma EC-1 cell line. Method: Inhibition of EC-1 cell proliferation at different concentrations of $As_20_3$ was assessed using the methyl thiazolyl blue colorimetric method (MTT method), with calculation of $IC_{50}$ value and choice of 20% of the $IC_{50}$ as the experimental drug concentration. Blank control, $As_20_3$, hyperthermia, radiotherapy group, $As_20_3$ + hyperthermia, $As_20_3$ + radiotherapy, hyperthermia + radiotherapy and $As_20_3$ + hyperthermia + radiotherapy groups were established, and the cell survival fraction (SF) was calculated from flat panel colony forming analysis, and fitted by the 'multitarget click mathematical model'. Flow cytometry (FCM) was used to detect changes in cell apoptosis and the cell cycle. Results: $As_20_3$ exerted inhibitory effects on proliferation of esophageal carcinoma EC-1 cells, with an $IC_{50}$ of 18.7 ${\mu}mol/L$. After joint therapy of $As_20_3$ + hyperthermia + radiotherapy, the results of FCM showed that cells could be arrested in the $G_2$/M phase, and as the ratio of cells in $G_0/G_1$ and S phases decreased, cell death became more pronounced. Conclusion: $As_20_3$ and hyperthermia exert radiosensitivity effects on esophageal carcinoma EC-1 cells, with synergy in combination. Mechanistically, $As_20_3$ and hyperthermia mainly influence the cell cycle distribution of EC-1 esophageal carcinoma cells, decreasing the repair of sublethal damage and inducing apoptosis, thereby enhancing the killing effects of radioactive rays.