Design, synthesis and biological evaluation of Thiazolo[3, 2-a]Pyrimidine derivatives as novel RNase H inhibitors.

Targeting Ribonuclease H (RNase H) has been considered a viable strategy for HIV therapy. In this study, a series of novel thiazolo[3, 2-a]pyrimidine derivatives were firstly designed and synthesized as potential inhibitors of HIV-1 RNase H. Among these compounds, A28 exhibited the most potent inhibition against HIV-1 RNase H with an IC50 value of 4.14 µM, which was about 5-fold increase in potency than the hit compound A1 (IC50 = 21.49 µM). To gain deeper insights into the structure-activity relationship (SAR), a CoMFA model was constructed to yield reasonable statistical results (q2 = 0.658 and R2 = 0.969). Results from magnesium ion chelation experiments and molecular docking studies revealed that these thiazolopyrimidine inhibitors may exert their inhibitory activity by binding to an allosteric site on RNase H at the interface between subunits p51 and p66. Furthermore, this analog demonstrated favorable physicochemical properties. Our findings provide valuable groundwork for further development of allosteric inhibitors targeting HIV-1 RNase H.

Theoretical Study on the Coordination and Separation Capacity of Macrocyclic N-Donor Extractants for Am(III)/Eu(III).

Designing ligands that can effectively separate actinide An(III)/lanthanide Ln(III) in the solvent extraction process remains one of the key issues in the treatment of accumulated spent nuclear fuel. Nitrogen donor ligands are considered as promising extractants for the separation of An(III) and Ln(III) due to their environmental friendliness. Four new macrocyclic N-donor hexadentate extractants were designed and their coordination with Am(III) and Eu(III), as well as their extraction selectivity and separation performance for Am(III) and Eu(III), were investigated by scalar relativistic density functional theory. A variety of theoretical methods have been used to evaluate the properties of the four ligands and the coordination structures, bonding properties, and thermodynamic properties of the complexes formed by the four ligands with Am(III) and Eu(III). The results of various wavefunction analysis methods including NBO analysis, quantum theory of atoms in molecules (QTAIM) analysis, and so on show that Am(III) has a stronger coordination ability with the ligands than Eu(III) due to the Am 5f orbitals more involved in bonding with the ligands than the Eu 4f orbitals, and the bonding environment of the N-donor in the ligand has a significant effect on its coordination ability of the metal ions. Thermodynamic analysis of the solvent extraction process shows that all of the four N-containing macrocyclic ligands have good extraction selectivity and separation performance for Am(III) and Eu(III). This study provides theoretical support for designing potential nitrogen-containing macrocyclic extractants with excellent separation performance.

Esketamine versus Sufentanil Applied Prior to Placement of Suspension Laryngoscope.

OBJECTIVE: In laryngeal microsurgery, the insertion of the suspension laryngoscope is a strong stimulus that may cause hemodynamic fluctuations and adverse cardiovascular events. The purpose of this study was to compare the effect of preemptive treatment with esketamine and sufentanil on maintaining hemodynamics and reducing the occurrence of adverse cardiovascular events during the insertion of suspension laryngoscope. METHODS: In this double-blind randomized controlled trial, patients undergoing general anesthesia for laryngeal microsurgery were randomly assigned (1:1) to esketamine 0.5 mg kg-1 (esketamine group) and sufentanyl 0.125 µg kg-1 (sufentanil group) before inserting the laryngoscope, respectively. RESULTS: During the insertion of suspension laryngoscope, the incidence of bradycardia (HR < 60 beats/min) was 39.3% (22/56) in esketamine group, lower than 60.0% (33/55) in sufentanil group (odds ratio [OR], 2.32 [95% CI, 1.11-5.08]; p = 0.029). The incidence of hypotension (MAP <65 mmHg) was 33.9% (19/56) in esketamine group, lower than 56.4% (31/55) in sufentanil group (odds ratio [OR], 2.52 [95% CI, 1.91-5.27]; p = 0.018). The frequency of hypotension in esketamine group was lower than that in sufentanil group (0.36 ± 0.52 vs. 0.56 ± 0.50, p = 0.035). The time-weighted average of HR dropping above 30% of baseline was smaller in esketamine group than in sufentanil group (0.52 ± 2.06 vs. 1.08 ± 2.77, p = 0.006). CONCLUSIONS: These findings showed that compared with preemptive treatment of sufentanil (0.125 µg kg-1 ), esketamine (0.5 mg kg-1 ) was effective in reducing the incidence of cardiovascular adverse events, including bradycardia and hypotension induced by the insertion of suspension laryngoscope during the laryngeal microsurgery. LEVEL OF EVIDENCE: 2 Laryngoscope, 133:3021-3027, 2023.

Applicability of Zipf's Law in Traditional Chinese Medicine Prescriptions.

Objective Traditional Chinese medicine (TCM) prescriptions have been used to cure diseases in China for thousands of years, in which many TCM herbs have no definite common quantity. Some key TCM herbs are commonly used and thus deserve in-depth investigations based on a more acceptable classification method. This study analyzes whether TCM prescriptions follow Zipf's law and attempts to obtain the thresholds of key TCM herbs based on the application of Zipf's law. Methods A total of 84,418 TCM prescriptions were collected and standardized. We tested whether Zipf's law and Zipf's distribution fit the Chinese herb distributions. A linear fitting experiment was performed to verify the relationship between the frequency distribution and frequency of TCM herbs. Results The distribution of TCM herbs in TCM prescriptions conformed to Zipf's law. Accordingly, the thresholds were obtained for the key TCM herbs. Conclusion The distribution of TCM herbs in TCM prescriptions follows Zipf's law.

[Identification of metabolites of Yiqi Baoyuan Prescription in rat plasma, bile, urine and feces after oral administration].

This study was designed to determine the metabolites of Yiqi Baoyuan Prescription(YQBYP) in rats. The ultra-high performance liquid chromatography coupled to time-of-flight mass spectrometry(UPLC-TOF-MS) and mass defect filter(MDF) were employed to analyze the metabolites of YQBYP in rat plasma, bile, urine and feces. Chromatographic separation was conducted on Acquity UPLC BEH C_(18) column(2.1 mm×100 mm, 1.7 µm) under gradient elution with 0.1% formic acid aqueous solution(A)-acetonitrile(B), and the column temperature was 30 ℃. Electrospray ion(ESI) source was used under positive and negative ion modes, with capillary voltage of 3.0 kV and mass scanning range of m/z 100-1 000. In this experiment, 9 prototype components and 36 metabolites were identified in rat plasma, bile, urine and feces samples. The results showed that the main metabolic pathways of YQBYP in rats involved methylation, demethylation, oxidation, and other phase Ⅰ reactions as well as glucuronidation, sulfation, and other phase Ⅱ reactions. This study provided scientific basis for clarifying the therapeutic material basis of YQBYP and product development.

[Determination of eight active components of Bufei Huoxue Capsules in rat plasma and their pharmacokinetics by UHPLC-MS/MS].

An ultra-high performance liquid chromatography-tandem mass spectrometry(UHPLC-MS/MS) method was established to investigate the pharmacokinetic behaviors of psoralenoside, isopsoralenoside, calycosin-7-glucoside, ononin, psoralen, isopsoralen, methylnissolin, and neobavaisoflavone in rat plasma after oral administration of Bufei Huoxue Capsules. After SD rats were administered with Bufei Huoxue Capsules suspension by gavage, blood samples were collected from the inner canthus at different time points. After protein precipitation, plasma samples were separated on ACQUITY UPLC BEH C_(18) column(2.1 mm×100 mm, 1.7 µm). The mobile phase consisted of acetonitrile(A) and water(B) containing 0.1% formic acid in gradient elution. The positive and negative ions were measured simultaneously in the multi-reaction monitoring(MRM) mode. The pharmacokinetic parameters were calculated and fitted by DAS 3.2.8. Psoralenoside, isopsoralenoside, calycosin-7-glucoside, ononin, psoralen, isopsoralen, methylnissolin, and neobavaisoflavone were detected in the rat plasma after drug administration, with AUC_(0-t) of(3 357±1 348),(3 555±1 696),(3.03±0.88),(2.21±0.33),(1 787±522),(2 295±539),(5.69±1.41) and(3.40±0.75) µg·L~(-1)·h, and T_(max) of(1.56±0.62),(1.40±0.70),(0.21±0.05),(0.25±0.12),(0.26±0.11),(0.34±0.29),(0.74±0.59), and 0.25 h. The method is proved specific and repeatable and is suitable for the determination of psoralenoside, isopsoralenoside, calycosin-7-glucoside, ononin, pso-ralen, isopsoralen, methylnissolin, and neobavaisoflavone in the rat plasma, which can be applied to pharmacokinetic study.

The distribution and epidemic characteristics of cerebrovascular disease in followed-up hypertension patients.

To explore distribution and epidemic characteristics of CVD in followed-up HP patients. Using the Hypertension Follow-up Management System database in Jiading district in Shanghai. We designed a retrospective cohort study that included all followed-up hypertension patients between 2002 and 2020. The endpoint was the occurrence of CVD confirmed by the hospital; otherwise, the patients were tracked until September 30, 2020. Record information of every patient has been collected in the registration card and each followed-up record. Among 223,097 observational followed-up HP patients, the total number of person years of observation was 4,244,421.25 person-year, 11,143 patients had developed CVD from hypertension before the deadline, the total incidence density was 0.00263 per person-year (male 0.00292; female 0.00238) and the complication ratio of CVD in HP patients was 4.99% (male 5.25%; female 4.76%) during follow-up period. The proportion of ischemic cerebrovascular, hemorrhagic cerebrovascular and unclassified stroke was respectively 71.18%, 5.95% and 22.87% in hypertensive CVD. Complication ratio of CVD increased with age, the group under 30 was 0, and the group over 70 was the highest (6.90%). The complication ratio of grad I, grad II and grad III blood pressure were respectively 4.79%, 4.96% and 6.13%. The complication ratio was 4.92% in only high systolic blood pressure patients; 17.23% in only high diastolic blood pressure patients; 4.59% in high systolic and diastolic blood pressure patients. The peak of complication ratio of CVD was 9-10 years after the registered and followed-up. The proportion of CVD cases in HP patients from April to June was the largest in the four seasons; the proportion of patients from October to December was the minimum. HP patient was prone to falling cerebrovascular disease; the main type of disease was cerebral infarction. Complication ratio in male incidence was higher than that in female. The complication ratio of CVD increased with age, blood pressure and duration of HP patients. It had seasonal characteristics, which was relatively high from April to June within year.

The interaction on hypertension between family history and diabetes and other risk factors.

To explore the individual effect and interaction of diabetes and family history and other risk factors on hypertension in Han in Shanghai China. The method of case-control study with l:l matched pairs was used, 342 cases of hypertension and 342 controls were selected and investigate their exposed factors with face-to-face. The method of epidemiology research was used to explore the individual effect and interaction of diabetes and family history and other risk factors on hypertension. The individual effect of family history (OR = 4.103, 95%CI 2.660-6.330), diabetes (OR = 4.219, 95%CI 2.926-6.083), personal taste (OR = 1.256, 95%CI 1.091-1.593), drinking behavior (OR = 1.391, 95%CI 1.010-1.914) and smoking behavior (OR = 1.057, 95%CI 1.00-1.117) were significant (p < 0.05). But individual effect of sex, education, occupation, work/life pressure, environmental noise, sleeping time and sports habit were not significant (p > 0.05). The OR of interaction between FH and DM to hypertension was 16.537 (95%CI 10.070-21.157), between FH and drinking behavior was 4.0 (95%CI 2.461-6.502), FH and sport habit was 7.668 (95%CI 3.598-16.344), FH and personal taste was 6.521 (95%CI 3.858-11.024), FH and smoking behavior was 5.526 (95%CI 3.404-8.972), FH and work/life pressure was 4.087 (95%CI 2.144-7.788). The SI of FH and DM was 2.27, RERI was 8.68, AP was 52.48% and PAP was 55.86%. FH and DM, personal taste, smoking behavior had positive interaction on hypertension, but FH and sport habits, drinking behavior, work/life pressure had reverse interaction on hypertension. FH and diabetes were very important risk factors with significant effect for hypertension. FH and diabetes, personal taste, smoking behavior had positive interaction on hypertension, but FH and sport habits, drinking behavior, work/life pressure had reverse interaction on hypertension.

Association Between Vitamin D and Hyperuricemia Among Adults in the United States.

Background: Serum uric acid can act as a risk factor for cardiovascular disease (CVD) and as antioxidant defense. Vitamin D deficiency can activate the parathyroid to induce the release of parathyroid hormone, which was thought to increase serum uric acid level, and low vitamin D status may also be associated with risk of CVD. No known studies have explored the association between serum 25(OH) D, vitamin D intake, and HU for the American population. Methods: We extracted 15,723 US adults aged 20-85 years from the National Health and Nutrition Examination Survey (NHANES) in 2007-2014. All dietary intakes were evaluated through 24-h dietary recalls. Multivariable logistic regression analysis was performed to examine the associations after adjustment for confounders. Results: Compared to the lowest quintile (Q1), for males, adjusted odds ratios (ORs) of HU in Q2 to Q4 of serum 25(OH) D levels were 0.78 (95% CI, 0.65-0.93), 0.97 (0.81-1.16), and 0.72 (0.60-0.88); ORs in Q2-Q5 of total vitamin D intake were 0.83 (0.69-0.98), 0.69 (0.58-0.83), 0.66 (0.55-0.79), and 0.59 (0.48-0.71), respectively. In females, OR was 0.80 (0.66-0.97) of serum 25(OH) D for Q3, and ORs in Q5 of total vitamin D intake were 0.80 (0.65-0.98). Conclusions: Our findings indicated that the serum 25(OH) D intakes of dietary vitamin D, supplemental vitamin D, and total vitamin D were inversely associated with HU in males. In females, a lower risk of HU with higher serum 25(OH) D, dietary vitamin D, and total vitamin D intake was found, but with no association between supplemental vitamin D intake and the risk of HU.

Nasal nebulization inhalation of budesonide for chronic rhinosinusitis with nasal polyps: A protocol for systematic review and meta-analysis.

BACKGROUND: Chronic rhinosinusitis and nasal polyps (CRNP) is a common public health concern for general population, and is thought to negatively impact their quality of life. Although previous studies have reported that nasal nebulization inhalation of budesonide (NNIB) can benefit patients with such condition, its conclusions are still inconsistent. Thus, this study will assess the efficacy and safety of NNIB for the treatment of CRNP. METHODS: To identify any associated studies, we will comprehensively and systematically search Cochrane Library, PubMed, EMBASE, Web of Science, PsycINFO, Cumulative Index to Nursing and Allied Health Literature, Allied and Complementary Medicine Database, Chinese Biomedical Literature Database, and China National Knowledge Infrastructure. We will search all electronic databases from inception to the present with no limitations of language and publication status. Two independent reviewers will undertake selection of study, data collection, and study quality evaluation, respectively. Another reviewer will help to settle down any different opinions between both of them. Study quality will be checked using Cochrane risk of bias tool, and statistical analysis will be performed using RevMan 5.3 software. RESULTS: This study will assess the efficacy and safety of NNIB for the treatment of CRNP through assessing primary outcomes of nasal symptoms and polyp sizes, and secondary outcomes of serum cortisol levels, health-related quality of life, and any expected and unexpected adverse events. CONCLUSION: The results of this study will summarize the up-to-date evidence on assessing the efficacy and safety of NNIB for the treatment of CRNP. STUDY REGISTRATION NUMBER: INPLASY202040108.

Effect of CO2 laser for the management of primary otosclerosis: A protocol for systematic review and meta-analysis.

BACKGROUND: This study will explore the effect and safety of CO2 laser (COL) for the management of patients with primary otosclerosis (PO). METHODS: The following electronic databases will be searched from inception to the present: PUBMED, EMBASE, The Cochrane Library, Web of Science, PsycINFO, Cumulative Index to Nursing and Allied Health Literature, Allied and Complementary Medicine Database, Chinese Biomedical Literature Database, VIP, WANGFANG, and China National Knowledge Infrastructure. No language limitation will be applied. All relevant randomized controlled trials using COL to treat patients with PO will be included. Two researchers will identify studies, collect data and evaluate the risk of bias of each included study independently. Any different views between 2 researchers will be resolved by a third researcher via discussion. Data analysis will be carried out using RevMan 5.3 software. RESULTS: This study will evaluate the effect and safety of COL for the treatment of PO through hearing gain, tinnitus severity, incidence of intraoperative, health-related quality of life, other morbidities, and adverse events. CONCLUSION: This study will provide evidence for the effect and safety of COL in patients with PO. STUDY REGISTRATION NUMBER: INPLASY202040110.

Efficacy of levocetirizine for the treatment of children with allergic rhinitis: A protocol for systematic review and meta-analysis.

BACKGROUND: Although previous studies have reported that levocetirizine is utilized for the treatment of children with allergic rhinitis (AR), its conclusions remain inconsistent. This study aims to evaluate the efficacy and harms of levocetirizine for children with AR. METHODS: Electronic database sources will be undertaken from the beginning to the present: MEDLINE, EMBASE, The Cochrane Library, CINAHL, ACMD, PsycINFO, Chinese Biomedical Literature Database, and China National Knowledge Infrastructure. We will not apply any restrictions to language and publication status. We will only consider randomized controlled trials of levocetirizine for children with AR. Two authors will independently scan literature, select studies, and collect data. Study quality for each included trial will be assessed using Cochrane risk of bias tool, and statistical analysis will be conducted using RevMan 5.3 software. RESULTS: This study will summarize the present evidence to systematically assess the efficacy and harms of levocetirizine for children with AR. CONCLUSION: The findings of this study intent to adequately inform stakeholders or clinicians, as well as to help develop treatment guidelines. STUDY REGISTRATION NUMBER: INPLASY202040111.

Familial aggregation and heritability of hypertension in Han population in Shanghai China: a case-control study.

BACKGROUND: To explore the familial aggregation and heritability of hypertension in Han in Shanghai China. METHODS: According to l:l matched pairs design, 342 patients of hypertension and 342 controls were selected and investigate their nuclear family members in the case-control study. The method of genetic epidemiology research was used to explore the familial aggregation and heritability of hypertension. RESULTS: The prevalence rate of hypertension of first-degree relatives was significantly higher (34.44%) than that of second- degree relatives (17.60%) and third-degree relatives (13.51%) in Han Population in Shanghai China. Separation ratio p was 0.217, and prevalence rate of case group relatives was higher than that of control group relatives. The results showed a phenomenon of familial aggregation in the distribution of hypertension. The heritability of first- degree relatives was 49.51%; that of second-degree relatives and third-degree relatives were respectively 23.42 and 21.41%. CONCLUSION: The distribution of essential hypertension has phenomenon of familial aggregation in Han Population in Shanghai China. The separation ratio of essential hypertension in this study shows that essential hypertension conform to the characteristics of multigene genetic disease. The heritability of first-degree relatives is bigger than that of second-degree relatives and third-degree relatives.

The effect of body mass index and its interaction with family history on hypertension: a case-control study.

BACKGROUND: BMI is an indicator commonly used in the world to measure the weight and height of the body, it reflects the comprehensive outcome of acquired lifestyle; FH is a sign reflecting the main role of genetic factors. This study aimed to evaluate the effect of BMI and interaction with FH on hypertension risk in Shanghai adult population. METHODS: According to l:l matched pairs design, 342 cases and 342 controls were selected and investigated in this study, this study was performed in Shanghai, China. Participants received face-to-face questionnaire survey, anthropometric tests and laboratory examinations. Relevant indicators that reflect obesity including BMI and waist to hip ratio (WHR) were calculated. Multivariate logistic regression analysis was applied to explore the association between factors and hypertension risk. Interactive effect was evaluated by synergy index (SI), relative excess risk due to interaction (RERI), attributable proportion due to interaction (AP) and the percentage of the interaction between the pure factors (PAP). RESULTS: Among 684 study participants aged 28-87 years old, the differences of mean age and height between case group and control group are no significant (p > 0.05), but the differences of mean of weight, WC, HC, BMI and WHR are significant (p < 0.001). The OR of FH on hypertension is 4.986 (95%CI: 2.832~ 8.877); the OR of BMI on hypertension is respectively: low weight is 1.528 (95%CI: 0.551~ 4.239), overweight is 3.333 (95%CI: 1.678~ 6.617) and obesity is 7.312 (95%CI: 3.556~ 15.035). The OR of interaction between FH and BMI to hypertension is 12.993 (95%CI: 7.426~22.734). SI is 1.90 (95% CI: 1.48~3.78), RERI is 5.67 (95% CI: 1.66~11.88), AP is 43.87% (95% CI: 12.84~91.88%), and PAP is 47.55% (95%CI: 13.91~99.58%). FH and BMI have positive interaction on hypertension. 43.87% of hypertension exposed to both FH and BMI was attributable to the interaction of them. CONCLUSIONS: FH and BMI are significant higher risks of hypertension; with the increase of BMI, the risk of hypertension will increase more. FH and BMI have positive interaction with hypertension, the interaction is greater than the sum of two independent actions.

Biomarkers of intravenous immunoglobulin resistance and coronary artery lesions in Kawasaki disease.

BACKGROUND: Currently, there are no reliable indicators for predicting intravenous immunoglobulin resistance and coronary artery lesions in the early stage of Kawasaki disease. METHODS: A total of 300 patients with Kawasaki disease were studied retrospectively. Laboratory data were compared between the intravenous immunoglobulin resistant (29 patients) and responsive groups, and between the groups with coronary artery lesions (48 patients) and without coronary artery lesions. RESULTS: The intravenous immunoglobulin resistant group had significantly higher D-dimer, globulin, interleukin-6 and serum ferritin levels in comparison to the intravenous immunoglobulin responder group. D-dimer level had a sensitivity of 87.0% and a specificity of 56.3% for predicting intravenous immunoglobulin resistance at a cutoff point of 1.09 mg/L. Globulin had a sensitivity of 62.1% and a specificity of 82.3% for predicting intravenous immunoglobulin resistance at a cutoff point of 34.7 g/L. Serum ferritin level had a sensitivity of 42.9% and a specificity of 88.8% for predicting intravenous immunoglobulin resistance at a cutoff point of 269.7 ng/mL. The patients with coronary artery lesions had higher D-dimer and tumor necrosis factor-α level. D-dimer level had a sensitivity of 50% and a specificity of 78.6% for predicting coronary artery lesions at a cutoff point of 1.84 mg/L. Based on analysis by multivariate logistic regression, serum ferritin and globulin were independent risks for intravenous immunoglobulin resistance, D-dimer was independent risk for coronary artery lesions. CONCLUSIONS: Elevated serum ferritin, globulin and D-dimer levels are significantly associated with intravenous immunoglobulin resistance in Kawasaki disease. Moreover, serum D-dimer is significantly increased in Kawasaki disease with coronary artery lesions.

Correlation between rs198388 and rs198389 polymorphismsin brainnatriuretic peptide (NPPB) gene and susceptibility to congenital heart diseases in a Chinese population.

OBJECTIVE: We discussed the correlation between SNP loci (rs198389 and rs198388) in brain natriuretic peptide gene (NPPB) and susceptibility to congenital heart diseases (CHD). METHOD: Multiplex SNaPshot technique was adopted for profiling of SNP genotypes at loci rs198389 and rs198388 in NPPB gene among 150 cases of CHDand 150 normal controls. RESULTS: The distribution frequency of 3 genotypes (AA, AG and GG) at locus rs198389 was 40.7%, 36.0% and 23.3% in CHD group, respectively, showing significant differences compared with the normal controls (P<0.001). Gallele was associated with higher risk of CHD (OR=2.48, 95% CI=1.77-3.48). The distribution frequency of CC, CTand TT genotypes at locus rs198388 was 60.7%, 17.3% and 22.0% in CHD group, respectively, also showing significant differences compared with the normal controls (P<0.001). C allele could increase the risk of CHD (OR=1.92, 95% CI=1.48-2.48). CONCLUSION: SNP loci rs198389 and rs198388 in NPPB gene were correlated with genetic susceptibility to CHD.

Co-expression of disulfide oxidoreductases DsbA/DsbC markedly enhanced soluble and functional expression of reteplase in Escherichia coli.

Reteplase is the third generation of thrombolytic medicine and has many advantages over commercial t-PA. However, over-expressing recombinant reteplase in E. coli always accumulates as inclusion bodies due to nine pairs of disulfide bonds formation that is the main obstacle for correct folding. In this paper, in order to enhance soluble expression of recombinant reteplase in E. coli, DsbA/DsbC foldases were used to introduce disulfide bonds into the reduced polypeptide chain and catalyze their isomerization to the native disulfide linkage during the folding process. Firstly multiple E. coli protein expression systems, i.e. DsbA, DsbC and DsbA/DsbC co-expression were constructed. Subsequently, IPTG and l-arabinose were added to induce expression of foldases and reteplase accordingly, and experimental parameters such as culture temperature and inducer concentration were optimized. As a result, the co-expression system markedly enhanced soluble expression of recombinant reteplase, and up to 60% of reteplase achieved soluble expression especially for the DsbC co-expression system. The fibrin plate method for active reteplase quantification showed that ∼70 mg soluble reteplase per liter fermentation broth was obtained with 2.35 × 105 IU/mg thrombolytic activity. Finally, fluorescence spectra indicated that the structural conformation of soluble reteplase was identical to its native state. The soluble expression of recombinant reteplase in E. coli was accomplished by co-expression with DsbA/DsbC, which contributes to further research in clinical application and folding mechanism, and provides guidance for production of other proteins with disulfide bonds.

Identification of metabolites of FR429, a potential antitumor ellagitannin, transformed by rat intestinal bacteria in vitro, based on liquid chromatography-ion trap-time of flight mass spectrometry analysis.

FR429 is an ellagitannin with a potential antitumor activity, isolated and purified from Polygonum capitatum Buch.-Ham.ex D.Don, which is a traditional Miao-nationality herbal medicine in Guizhou and Yunnan of China. Our preliminary result of pharmacology study has indicated that the antitumor activity of FR429. However, the metabolism of FR429 has not been reported yet. In this study, LC-ion trap-time of flight mass spectrometry (LC-IT-TOF/MS) was used to characterize unpredictable metabolites of FR429 biotransformed by intestinal bacteria in vitro. Total thirteen metabolites were detected and characterized via comparisons of their accurate molecular masses and fragment ions of each MS(n) stage with those of the parent drug, and four of them were also elucidated by NMR. The results demonstrated that FR429 could be transformed by intestinal bacteria in vitro, mainly via hydrolysis and reduction reaction. This work provided a basis for the further study on the biotansformation of FR429 in vivo.

In vitro study on metabolite profiles of bioactive xanthones isolated from Halenia elliptica D. Don by high performance liquid chromatography coupled to ion trap time-of-flight mass spectrometry.

The metabolisms of five xanthones isolated from a Tibetan medicinal herb Halenia elliptica D. Don, including 1-hydroxy-2,3,5-trimethoxy-xanthone (HM-1), 1-hydroxy-2,3,4,7-tetramethoxy-xanthone (HM-2), 1-hydroxy-2,3,4,5-tetramethoxy-xanthone (HM-3), 1,7-dihydroxy-2,3,4,5-tetramethoxy-xanthone (HM-4) and 1,5-dihydroxy-2,3-dimethoxy-xanthone (HM-5), were studied in rat liver microsomes in vitro. High performance liquid chromatography coupled to ion trap time-of-flight mass spectrometry (LC-ESI-IT-TOF) was applied for identification of metabolites of five xanthones mentioned above and (1)H NMR was used to elucidate the major metabolites. The structures of thirteen metabolites were identified and seven of them had not been reported before. Moreover, xanthone isomers herein could be distinguished by difference of fragmentation behaviors with increase of stages or relative abundances. The results indicated that in vitro metabolic transformation of HM-1, HM-2, HM-3, HM-4 and HM-5 occurred mainly at 2-, 4-, 5-, 7-carbonic positions on their structures of parent drugs. The metabolites could be new vasoactive substances. This work will provide a basis for study on the structure-activity relationships of these xanthones and their derivatives from Tibetan herbal in the next work.

High performance liquid chromatographic separation of thirteen drugs collected in Chinese Pharmacopoeia 2010(Ch.P2010) on cellulose ramification chiral stationary phase.

The enantiomers separation of thirteen drugs collected in Ch.P2010 was performed on chiral stationary phase of cellulose ramification (chiralpak OD and chiralpak OJ) by high performance liquid chromatographic (HPLC) methods, which included ibuprofen (C1), ketoprofen (C2), nitrendipine (C3), nimodipine (C4), felodipine (C5), omeprazole (C6), praziquantel (C7), propranolol hydrochloride (C8), atenolol (C9), sulpiride (C10), clenbuterol hydrochloride (C11), verapamil hydrochloride (C12), and chlorphenamine maleate (C13). The mobile phase consisted of isopropanol and n-hexane. The detection wavelength was set at 254 nm and the flow rate was 0.7 mL/min. The enantiomers separation of these thirteen racemates on chiralpak OD column and chiralpak OJ column was studied, while the effects of proportion of organic additives, alcohol displacer and temperature on the separation were studied. And the mechanism of some of racemates was discussed. The results indicated that thirteen chiral drugs could be separated on chiral stationary phase of cellulose ramification in normal phase chromatographic system. The chromatographic retention and resolution of enantiomers could be adjusted by factors including column temperature and the concentration of alcohol displacer and organic alkaline modifier in mobile phase. It was shown that the resolution was improved with reducing concentration of alcohol displacer. When concentration of organic alkaline modifier was 0.2% (v/v), the resolution and the peak shape were fairly good. Most racemates mentioned above had better resolution at column temperature of 25 °C. When racemates were separated, the temperature should be kept so as to obtain stable separation results.