Modification of nitrilase based on computer screening and efficient biosynthesis of 4-cyanobenzoic acid.

4-cyanobenzoic acid serves as a crucial intermediate for the synthesis of various high-value organic compounds. The enzymatic hydrolysis of terephthalonitrile to produce 4-cyanobenzoic acid using nitrilase offers the advantages of a simple reaction pathway, environmental friendliness, and easy product separation. In order to efficiently develop nitrilases that meet industrial production requirements, the virtual screening method used in the study is established and mature. From a total of 371 amino acids in the nitrilase AfNIT, which exhibits activity in terephthalonitrile hydrolysis, three candidate sites (F168, S192, and T201) were identified, and a "small and accurate" mutant library was constructed. The triple mutant F168V/T201N/S192F was screened from this small mutant library with a specific activity of 227.3 U mg-1 , which was 3.8 times higher than that of the wild-type AfNIT. Using the whole-cell biocatalyst containing the mutant F168V/T201N/S192F, terephthalonitrile was successfully hydrolyzed at a concentration of 150 g L-1 to produce 4-cyanobenzoic acid with a final yield of 170.3 g L-1 and a conversion rate of 98.7%. The obtained nitrilase mutant F168V/T201N/S192F in this study can be effectively applied in the biomanufacturing of 4-cyanobenzoic acid using terephthalonitrile as a substrate. Furthermore, the results also demonstrate the significant improvement in predictive accuracy achieved through the latest AI-assisted computer simulation methods. This approach represents a promising and feasible new technological pathway for assisting enzyme engineering research, laying a theoretical foundation for other related studies.

Doxorubicin induces stemness of mouse TNBC 4T1 cells through Stat3-Oct-4/CD44 signaling pathway / 中国病理生理杂志

AIM:To investigate the stemness of mouse triple-negative breast cancer (TNBC) 4T1 cells induced by doxorubicin (DOX) and the underlying mechanism.METHODS:The 4T1 cells and MDA-MB-468 cells were treated with DOX at different concentrations (0, 0.05, 0.1 and 0.5μmol/L) for 24 h, and the shape and viability of the cells were observed.The concentration of DOX at 0.1μmol/L was chosen as the optimal concentration for the following experiments.The 4T1 cells and MDA-MB-468 cells resistant to DOX were established by continuous stimulation with DOX for 4 weeks, and named as 4T1-DOX and MDA-MB-468-DOX.Sphere formation assay was used to detect the stemness of 4T1cells and MDA-MB-468 cells.The expression of CD133 was observed by immunofluorescence staining.The expression of CD44 was analyzed by flow cytometry.The protein levels of Stat3, phosphorylated Stat3 (p-Stat3) and Oct-4 were determined by Western blot.RESULTS:The sphere formation ability of the 4T1-DOX cells was stronger than that of the 4T1control cells.The 4T1-DOX cells expressed high levels of the stemness markers CD133 and CD44 as compared with the 4 T1 cells (P<0.05).Furthermore, the 4T1-DOX cells exhibited enhanced activation of Stat3 (p-Stat3) and increased expression of Oct-4 (P<0.05) , while the expression of total Stat3 had no obvious variation.In addition, when activation of Stat3 was inhibited by WP1066, the protein levels of p-Stat3, Oct-4 and CD44 were down-regulated (P<0.05).Furthermore, inhibition of Stat3 phosphorylation reduced the sphere formation ability of the 4T1-DOX cells (P<0.05).CONCLUSION:DOX induces the stemness of mouse TNBC 4T1 cells through Stat3-Oct-4 signaling pathway.

Detection of fluoride ion concentration in osteoblasts in vitro by nuclear magnetic resonance spectroscopy / 中国地方病学杂志

ObjectiveTo detect the concentration and distribution of fluoride ions in osteoblasts exposed to fluoride in vitro culture,and to provide basic information for studying the effect of fluoride on osteoblast injury.MethodsIn vitro cultured osteoblasts were exposed to 0,5,10,20,40 mg/L fluoride for 3,10,30 d (n =6),respectively.Concentration and distribution of fluoride ions in the cytoplasm and the nucleus of these osteoblasts were determined by nuclear magnetic resonance spectroscopy.Results(①) After cultured for 3 d,fluoride ion content of the bone cytoplasm exposed to different concentrations of fluoride 0,5,10,20,40 mg/L were (0.83 ±0.65),(0.54 ± 0.23),(0.65 ± 0.77),(0.59 ± 0.87),(3.64 ± 1.21 )mg/L,respectively,and the values of exposed to 40 mg/L fluoride group was significantly higher than that of exposed to 0,5 mg/L groups (all P ＜ 0.05).(②)after cultured for 10 d,the composition of the fluoride ion in cytoplasm of exposed to fluoride 10,20,40 mg/L groups were (4.03 ± 1.23),(3.66 ± 0.98),(6.26 ± 2.10)mg/L,respectively,which were higher than that of exposed to 0,5 mg/L groups [(0.78 ± 0.75),(2.69 ± 0.89)mg/L,respectively,all P ＜ 0.05].Of fluoride 20,40 mg/L groups,the composition of the fluoride ion in nucleus were (1.63 ± 1.19),(2.17 ± 1.21 )mg/L,respectively,which were higher than that of 0,5 mg/L groups[(0.65 ± 0.46),(1.57 ± 0.33) mg/L,all P ＜ 0.05].(③)After cultured for 30 d,of the exposed to fluoride 10,20,40 mg/L groups,the composition of the fluoride ion in cytoplasm were (3.99 ± 0.84),(4.33 ± 1.67),(5.80 ± 1.38)mg/L,respectively,which were higher than that of 0,5 mg/L groups[(0.88 ± 0.44),(2.84 ± 0.43)mg/L,all P ＜ 0.05].The composition of the fluoride ion in nucleus of the fluoride 20,40 mg/Lgroups were (3.33 ± 1.46),(3.53 ± 1.22)mg/L,respectively,which were significantly higher than that of 0,5mg/L groups [(0.70 ± 0.66),(1.99 ± 0.76)mg/L,all P ＜ 0.05].ConclusionsWhen osteoblasts are exposed to fluoride environment,fluoride ions enter into the osteoblasts quickly,and quickly accumulate in the nucleus,showing a special affinity between fluoride and bone tissue.Intracellular fluoride ions increase with the increase of contact time and exposure dose.

Effect of submarine training on antioxidant ability in submarine men / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To study the effect of the submarine training on the antioxidant ability of the submarine men.</p><p><b>METHODS</b>50 sea-training submarine men, 50 land-training submarine men and 50 resting submarine men were randomly selected from some submarine troops. The blood routine, the total antioxidative capacity (T-AOC), the content of malondialdehyde (MDA) and the levels of IFN-gamma in blood plasma, the hemolytic degree of RBC, the proliferation of peripheral-blood lymphocyte (PPL) of them were detected in each group.</p><p><b>RESULTS</b>The T-AOC of the sea-training submarine men, the land-training submarine men and the resting submarine men significantly increased by turns [(15.38 +/- 3.11), (18.81 +/- 2.45), (20.93 +/- 2.95) U/ml], but MDA and the hemolytic degree of RBC significantly decreased by turns [(2.56 +/- 0.70), (2.12 +/- 0.53),(1.77 +/- 0.56) nmol/ml and 25.72% +/- 1.67%, 21.45% +/- 1.02%, 18.28% +/- 1.37%] (P < 0.05). Compared with the land-training submarine men and the resting submarine men, IFN-gamma [(31.89 +/- 3.52) pg/ml] and the proliferation of PPL of the sea-training submarine men were significantly lower, whereas the red blood count (RBC) and hemoglobin (Hb) were significantly higher (P < 0.05).</p><p><b>CONCLUSION</b>Submarine training, especially sea training, may decrease the antioxidant ability.</p>

Nutrition protective research on oxidative damage to submarine men / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To investigate the effect of nutrition protection on oxidation damage of the submarine men.</p><p><b>METHODS</b>50 submarine men were randomly divided into test group and control group, 25 persons each. The test group member took VitB2 5 mg, VitC 200 mg, GPC capsule 50 mg, once every other day and VitA capsules 25 000 units for every week during the sea-voyage. The total anti-oxidative capacity (T-AOC), superoxide dismutase (SOD), malondialdehyde (MDA), the proliferation of peripheral-blood lymphocyte (PPL), the hemolytic degree of RBC and IFN-gamma were detected.</p><p><b>RESULTS</b>Before sea voyage, the difference in the T-AOC and SOD and PPL and IFN-gamma and the MDA content and the hemolytic degree of RBC between the test group and the control group were not significant (P>0.05). After sea voyage, the T-AOC and SOD and PPL and IFN-gamma in the test group [(24.08 +/- 0.10) U/ml, (44.85 +/- 0.96) U/ml, (0.29 +/- 0.05) (with H2O2), (0.34 +/- 0.04) (without H2O2) and (34.21 +/- 3.52) pg/ml] were higher than the control group [(21.06 +/- 1.10) U/ml, (42.80 +/- 1.46) nu/ml, (0.23 +/- 0.01) (with H2O2), (0.34 +/- 0.04) (without H2O2) and (31.89 +/- 3.52) pg/ml]. The MDA content and the hemolytic degree of RBC [(2.15 +/- 0.28) nmol/ml and (20.96% +/- 0.10%)] were lower than the control group [(2.44 +/- 0.32) nmol/ml and (23.12% +/- 0.77%)]. The difference was significant (P<0.05).</p><p><b>CONCLUSION</b>To add antioxidant nutrients can improve the submarine men's antioxidant capacity.</p>

Study on protective effect of grape procyanidins in radiation injury in radiation-contacted persons / 中华预防医学杂志

<p><b>OBJECTIVE</b>To study the protective effect of Grape procyanidins (GPC) on radiation injury in radiation-contacted persons.</p><p><b>METHODS</b>Sixty radiation-contacted persons were randomly divided into the experimental group and the control group and 15 radiation-uncontacted persons were selected as the normal group. The experimental group was given GPC (100 mg/day), while the control group was given the capsule of starch every day for 60 days. Vein blood samples were taken before and after the study and the total antioxidative capacity (T-AOC), Malondialdehyde (MDA), cell proliferation, expression levels of proliferative cell nuclear antigen (PCNA), Bcl-2 and Bax protein, WBC were measured.</p><p><b>RESULTS</b>The WBC, T-AOC and cell proliferation rate of the experimental group were (5.62 +/- 0.40) 10(9)/L, (17.07 +/- 1.91) U/ml and 0.87 +/- 0.09 respectively, which were significantly higher than those of the control group. The MDA and Bax expression levels were (4.12 +/- 0.37) nmol/L and 28.06% +/- 5.79% respectively that were significantly lower than those of the control group.</p><p><b>CONCLUSION</b>GPC should have protective effects on radiation injury of the radiation-contacted persons.</p>

Analysis of correlations between blood lead levels and calcium, iron, zinc and environmental factors in children / 中国地方病学杂志

Objective To investigate the current status of lead(Pb) poisoning in children and correlations between Pb levels and calcium(Ca), iron(Fe), zinc(Zn) and environmental factors. Methods ①Questionnaire survey was conducted among 3160 parents in Qingdao area, regarding factors that might have effects on the levels of Pb. ②Levels of blood Pb, Ca, Fe and Zn were determined by graphite furnace atomic absorption (GFAA) spectrometry for 3160 children, who were divided into three groups: mildly increased Pb levels, moderately increased Pb levels and severely increased Pb levels, and the effects of Ca, Fe and Zn on children's blood Pb levels were analyzed by multiple linear regressions.③Children having mildly Pb increase were divided into four groups and received Ca, Fe or Zn preparations or placebo. Results ① Children living with smokers, in new houses, in houses or kindergartens located in industrial areas or main traffic roads, consuming preserved eggs or canned drinks regularly were more easily poisoned.②Percentages of children with increased levels who had insufficient blood Ca, Fe and Zn : 53.31% (306/574), 46.17% (265/574) and 68.47% (393/574), respectively, in the group of children with mildly increased Pb levels; 57.68% (263/456), 49.34% (225/456) and 77.41% (353/456), respectively, in the group of children with moderately increased Pb levels; 64.71% (11/17), 58.82% (10/17) and 82.35%(14/17), respectively, in the group of children with severely increased Pb levels. In contrast, the percentages of children in the control group who had low blood Ca, Fe and Zn were 19.78%(418/2113), 5.21% (110/2113) and 14.62%(309/2113), respectively. ③The levels of Ca, Fe and Zn had a negative correlations with blood levels of Pb(coefficient of regression were-0.870,-0.430,-0.565, respectively). ④When given Ca, Fe and Zn, the children had the blood Pb levels decreased by 78.32% (112/143), 66.43% (95/143) and 67.83% (97/143), respectively. Conclusions ①Children's increased levels of Pb have adverse effects on the absorption of Ca, Fe and Zn. ②Intaking Ca, Fe and Zn preparations can reduce blood Pb level.

Inhibitory effect of grape procyanidin on the cell apoptosis induced by radiation / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To study the inhibition effect of grape procyanidin (GPC) on the cell apoptosis and injury of proliferation induced by radiation.</p><p><b>METHODS</b>Three indices including apoptosis rate, proliferation rate and expression of bcl-2 and bax protein were examined in the mice pancreas after taking different dose GPC by mouth and radiation by (60)Co-gamma ray once.</p><p><b>RESULTS</b>The cell proliferation and bcl-2 expression in high dose GPC group (3.16% +/- 0.13% and 49.8% respectively), were higher than those in radiation control group (0.64% +/- 0.11%, 29.7%), but the cell apoptosis rate and bax expression (19.8% and 55.0% respectively), were lower than those in radiation control group (35.6%, 85.7%). All the above differences were statistically significant (P < 0.05).</p><p><b>CONCLUSION</b>GPC has certain protective effect against the mice pancreatic cell apoptosis and the abnormal expression of bcl-2 and bax protein induced by (60)Co-gamma.</p>

Application of organic polymeric flocculants in centrifugal dewatering of oil refinery sludge.

In order to evaluate the applicability of the organic polymeric flocculants (OPF) in the treatment of oil refinery sludge, experiments were conducted to show that OPF have better performance of flocculation than inorganic flocculants. Both the anionic and cationic OPF have satisfactory flocculation efficiency in oil sludge treatment, but the latter are more cost-efficient. Among the over 20 types of flocculants tested, 2 OPF (CPAM-2 and HPAM-2) were selected as the treatment agents, based on their good treatment performances, oil-resistance and economic feasibility. It was demonstrated in the industrial-scale centrifugal dewatering experiments that the application of either CPAM-2 or HPAM-2 could achieve high treatment efficiency of the oil sludge dewatering and reduce the COD of centrifugal liquid to less than 1000 mg/L.

Micro-electrolysis technology for industrial wastewater treatment.

Experiments were conducted to study the role of micro-electrolysis in removing chromaticity and COD and improving the biodegradability of wastewater from pharmaceutical, dye-printing and papermaking plants. Results showed that the use of micro-electrolysis technology could remove more than 90% of chromaticity and more than 50% of COD and greatly improved the biodegradability of pharmaceutical wastewater. Lower initial pH could be advantageous to the removal of chromaticity. A retention time of 30 minutes was recommended for the process design of micro-electrolysis. For the use of micro-electrolysis in treatment of dye-printing wastewater, the removal rates of both chromaticity and COD were increased from neutral condition to acid condition for disperse blue wastewater; more than 90% of chromaticity and more than 50% of COD could be removed in neutral condition for vital red wastewater.