Objective To evaluate the promotional effectiveness of different TCM appropriate technology packages; To provide a scientific basis for improving the technology package. Methods 8 assessment indexes, including input costs, the number of experts, the number of people with senior professional post, the number of grassroots people under the guidance of experts, the number of training hours, the number of people receiving training, evaluating rate and the number of users were selected. TOPSIS method was used to evaluate the promotional effectiveness of five kinds of TCM packages in grassroots. Results The Ci values for appropriate technology packages were 0.76 for pains in neck, shoulder, waist and lower extremities, 0.49 for gynecological diseases, 0.44 for infantile diarrhea, 0.66 for chronic gastropathy, and 0.00 for tumors. Conclusion The effectiveness of promotion of TCM appropriate technology packages from high to low is pains in neck, shoulder, waist and lower extremities package, chronic gastropathy package, gynecological diseases package, infantile diarrhea package, tumors package.

The spices, with a wide range of remarkable pharmacological effects, were limited in developing clinical drugs because of their poor solubility, high irritation and low bioavailability. Controlled release preparations can delay the release of drugs on the basis of solubility enhancement to improve bioavailability. The research on spice ingredient-loaded controlled release preparations was sum-marized in the paper for the further development of spice active ingredients.

The main active components in Ginkgo biloba extracts were Ginkgo biloba flavonoids and lactone compounds. This pa-per reviewed on the kinds and pharmacological effects of the active ingredients in Ginkgo biloba extracts, and focused on four aspects including controlled-release preparations, solubilized solid preparations, nanoparticle formulations and time- and site-specific formula-tions to introduce the development in the new dosage forms of Ginkgo biloba flavonoids and lactone compounds.

Semen Ziziphi spinosae ( SZS) , as a commonly used traditional Chinese medicine in clinic for insomnia therapy, is rich in pharmacological active ingredients such as saponins, flavonoids, alkaloids, oil and the other chemical compounds. The recent stud-ies indicated that some active ingredients in SZS exhibited a variety of activities including sedative hypnotics, antianxiety and anti-de-pression by regulating particular neurotransmitters such as 5-hydroxytryptamine (5-HT), gamma-amino butyric acid (GABA), norepi-nephrine, dopamine and glutamate. In accordance with the previous studies on pharmacological activities of SZS, this paper summa-rized and reviewed the applications of SZS in the treatment of central nervous system diseases, myocardial diseases and hepatic disea-ses, which might provide solid evidence for the application development of SZS.

Aim To investigate the intracellular disposition process of doxorubicin (DOX) in human breast cancer MCF-7, providing reference for explaining the pharmacology and their side effects of anti-tumor drugs. Methods The drug-resistant cell line MCF-7/DOX of breast cancer with DOX indication was selected as the material, and ultra-high performance liquid chromatography quadrupole tandem time-of-flight high-resolution mass spectrometry (UPLC-Q-TOF-MS/MS) method was established to analyze the disposal of DOX by target cells. Results Two unreported trace a-mounts of new metabolites of doxorubicin were found, and their structures were deduced by high-resolution multistage mass spectrometry. Molecular docking showed that its affinity for DNA was lower than that of DOX. Conclusion Target cells have unique and diverse drug metabolism pathways for DOX, which may be related to drug resistance mechanisms.

Aim To investigate the effect of vaccarin on mouse atherosclerosis in vivo and the underlying mechanism. Methods AopE mice aged 6 to 8 weeks old were used to establish the atherosclerosis model. Oil red O staining was used to determine the lipid levels in aorta and aortic root. Enzyme linked immunosorbent assay (ELISA) was used to detect the levels of serum inflammatory factors. Results Vaccarin could effectively reduce the levels of blood glucose and blood pressure in AopE

The influence of different affinity tags on enzyme characteristics varies. The (S)-carbonyl reductase 2 (SCR2) from Candida parapsilosis can reduce 2-hydroxyacetophenone, which is a valuable prochiral ketones. Different affinity tags, i.e. his-tag, strep-tag and MBP-tag, were attached to the N terminus of SCR2. These tagged SCR2 enzymes, i.e. his6-SCR2, strep-SCR2 and MBP-SCR2, were heterologously expressed in Escherichia coli and purified to study their characteristics towards 2-hydroxyacetophenone reduction. Affinity tags did affect the characteristics of the recombinant SCR2 enzymes. Specifically, affinity tags affect the stability of recombinant SCR2 enzymes: 1) At pH 6.0, the remaining enzyme activities of his6-SCR2 and strep-SCR2 were only 95.2% and 90.0% of the untagged SCR2, while that of MBP-SCR2 was 1.2 times of the untagged SCR2 after incubating for 13 h at 30 °C. 2) The half-life of MBP-SCR2 at 50 °C was 26.6%-48.8% longer than those of strep-SCR2, his6-SCR2 and untagged SCR2. 3) The kcat of MBP-SCR2 was about 1.25-1.45 times of that of small affinity-tagged and untagged SCR2 after storing at -80 °C for 60 d. Structural informatics indicated that the α-helices at the C terminus of MBP-SCR2 contributed to the stability of the N terminus of fusion protein of SCR2. Data from circular dichroism showed that the MBP-tag has some influence on the secondary structure of SCR2, while melting temperature analysis demonstrated that the Tm of the recombinant MBP-SCR2 was about 5 °C higher than that of the untagged SCR2. This study obtained an efficient and stable recombinant SCR2, i.e. the MBP-SCR2. Moreover, this study could serve as a reference for other researchers to evaluate and select appropriate affinity tags for their research.
Alcohol Oxidoreductases ; Escherichia coli/genetics* ; Recombinant Fusion Proteins/genetics*

A whole-cell catalyst using Escherichia coli BL21(DE3) as a host, expressing L- threonine dehydratase from Escherichia coli, and co-expressing leucine dehydrogenase from Bacillus cereus and glucose dehydrogenase from Bacillus subtilis for cofactor regeneration, was constructed and used for one-pot production of L-2-aminobutyric acid (L-ABA) and D- gluconic acid from L-threonine and D-glucose. We used shake-flask culture to study the whole-cell catalytic condition including temperature, pH, proper permeabilization of cells and optimal wet cells amount. Moreover, the whole-cell catalyst was cultured in 5-L fermentor by fed-batch fermentation, and 164 g/L L-threonine and 248 g/L D-glucose were converted to 141.6 g/L L-ABA and 269.4 g/L D-gluconic acid. The whole-cell catalyst is promising to fulfill industrial requirements for L-ABA and D-gluconic acid.

2-Hydroxybutyric acid (2-HBA) is an important intermediate for synthesizing biodegradable materials and various medicines. Chemically synthesized racemized 2-HBA requires deracemization to obtain optically pure enantiomers for industrial application. In this study, we designed a cascade biosynthesis system in Escherichia coli BL21 by coexpressing L-threonine deaminase (TD), NAD-dependent L-lactate dehydrogenase (LDH) and formate dehydrogenase (FDH) for production of optically pure (S)-2-HBA from bulk chemical L-threonine (L-Thr). To coordinate the production rate and the consumption rate of the intermediate 2-oxobutyric acid in the multi-enzyme cascade catalytic reactions, we explored promoter engineering to regulate the expression levels of TD and FDH, and developed a recombinant strain P21285FDH-T7V7827 with a tunable system to achieve a coordinated multi-enzyme expression. The recombinant strain P21285FDH-T7V7827 was able to efficiently produce (S)-2-HBA with the highest titer of 143 g/L and a molar yield of 97% achieved within 16 hours. This titer was approximately 1.83 times than that of the highest yield reported to date, showing great potential for industrial application. Our results indicated that constructing a multi-enzyme-coordinated expression system in a single cell significantly contributed to the biosynthesis of hydroxyl acids.
Escherichia coli/genetics* ; Formate Dehydrogenases ; Hydroxybutyrates ; Threonine Dehydratase

Glutamic acid is an important amino acid with wide range of applications and huge market demand. Therefore, by performing transcriptome sequencing and re-sequencing analysis on Corynebacterium glutamicum E01 and high glutamate-producing strain C. glutamicum G01, we identified and selected genes with significant differences in transcription and gene levels in the central metabolic pathway that may have greatly influenced glutamate synthesis and further increased glutamic acid yield. The oxaloacetate node and α-ketoglutarate node play an important role in glutamate synthesis. The oxaloacetate node and α-ketoglutarate node were studied to explore effect on glutamate production. Based on the integrated strain constructed from the above experimental results, the growth rate in a 5-L fermenter was slightly lower than that of the original strain, but the glutamic acid yield after 48 h reached (136.1±5.53) g/L, higher than the original strain (93.53±4.52) g/L, an increase by 45.5%; sugar-acid conversion rate reached 58.9%, an increase of 13.7% compared to 45.2% of the original strain. The application of the above experimental strategy improved the glutamic acid yield and the sugar-acid conversion rate, and provided a theoretical basis for the metabolic engineering of Corynebacterium glutamicum.
Citric Acid Cycle ; Corynebacterium glutamicum/metabolism* ; Glutamic Acid/metabolism* ; Metabolic Engineering ; Metabolic Networks and Pathways/genetics*