[Main components from cultivated and wild Nardostachyos Radix et Rhizoma by LC-MS and GC-MS].

In this study, ultra-performance liquid chromatography-quadrupole/time-of-flight mass spectrometry(UPLC-Q-TOF-MS) and gas chromatography-mass spectrometry(GC-MS) were combined with non-targeted metabonomic analysis based on multivariate statistics analysis, and the content of five indicative components in nardosinone was determined and compared by UPLC. The main chemical components of Nardostachyos Radix et Rhizoma with imitative wild cultivation and wild Nardostachyos Radix et Rhizoma were comprehensively analyzed. The results of multivariate statistical analysis based on liquid chromatography-mass spectrometry(LC-MS) and GC-MS were consistent. G1 and G2 of the imitative wild cultivation group and G8-G19 of the wild group were clustered into category 1, while G7 of the wild group and G3-G6 of the imitative wild cultivation group were clustered into category 2. After removing the outlier data of G1, G2, and G7, G3-G6 of the imitative wild cultivation group were clustered into one category, and G8-G19 of the wild group were clustered into the other category. Twenty-six chemical components were identified according to the positive and negative ion modes detected by LC-MS. The content of five indicative components(VIP>1.5) was determined using UPLC, revealing that chlorogenic acid, isochlorogenic acid A, isochlorogenic acid C, linarin, nardosinone, and total content in the imitative wild cultivation group were 1.85, 1.52, 1.26, 0.90, 2.93, and 2.56 times those in the wild group, respectively. OPLS-DA based on GC-MS obtained 10 diffe-rential peaks. Among them, the relative content of α-humulene and aristolene in the imitative wild cultivation group were extremely significantly(P<0.01) and significantly(P<0.05) higher than that in the wild group, while the relative content of 7 components such as 5,6-epoxy-3-hydroxy-7-megastigmen-9-one, γ-eudesmol, and juniper camphor and 12-isopropyl-1,5,9-trimethyl-4,8,13-cyclotetrade-catriene-1,3-diol was extremely significantly(P<0.01) and significantly(P<0.05) lower than that in the wild group, respectively. Therefore, the main chemical components of the imitative wild cultivation group and wild group were basically the same. However, the content of non-volatile components in the imitative wild cultivation group was higher than that in the wild group, and the content of some volatile components was opposite. This study provides scientific data for the comprehensive evaluation of the quality of Nardostachyos Radix et Rhizoma with imitative wild cultivation and wild Nardostachyos Radix et Rhizoma.

Human induced pluripotent stem cell (hiPSC)-derived cardiomyocyte modelling of cardiovascular diseases for natural compound discovery.

Cardiovascular disease (CVD) remains the leading cause of death worldwide. Natural compounds extracted from medicinal plants characterized by diverse biological activities and low toxicity or side effects, are increasingly taking center stage in the search for new drugs. Currently, preclinical evaluation of natural products relies mainly on the use of immortalized cell lines of human origin or animal models. Increasing evidence indicates that cardiomyopathy models based on immortalized cell lines do not recapitulate pathogenic phenotypes accurately and a substantial physiological discrepancy between animals and humans casts doubt on the clinical relevance of animal models for these studies. The newly developed human induced pluripotent stem cell (hiPSC) technology in combination with highly-efficient cardiomyocyte differentiation methods provides an ideal tool for modeling human cardiomyopathies in vitro. Screening of drugs, especially screening of natural products, based on these models has been widely used and has shown that evaluation in such models can recapitulate important aspects of the physiological properties of drugs. The purpose of this review is to provide information on the latest developments in this area of research and to help researchers perform screening of natural products using the hiPSC-CM platform.

[Comparison of odor and quality of Galli Gigerii Endothelium Corneum derived from domestic chickens and broilers].

Galli Gigerii Endothelium Corneum(GGEC) is commonly used for the clinical treatment of indigestion, vomiting, diarrhea, and infantile malnutrition with accumulation. In recent decades, omnivorous domestic chickens, the original source of GGEC, has been replaced by broilers, which may lead to significant changes in the quality of the yielding GGEC. Through subjective and objective sensory evaluation, biological evaluation, and chemical analysis, this study compared the odor and quality between GGEC derived from domestic chickens and that from broilers. The odor intensity between them was compared by odor profile analysis and it was found that the fishy odor of GGEC derived from domestic chickens was significantly weaker than that of GGEC from broilers. Headspace-solid phase microextraction-gas chromatography-triple quadrupole tandem mass spectrometry(HS-SPME/GC-QQQ-MS/MS) suggested that the overall odor-causing chemicals were consistent with the fishy odor-causing chemicals. According to the odor activity va-lue and the orthogonal partial least squares discriminant analysis(OPLS-DA) result, dimethyl trisulfide, 2-methoxy-3-isobutylpyrazine, and 2-methylisoborneol were responsible for the fishy odor(OAV≥1) and the content of fishy odor-causing chemicals in GGEC derived from broilers was 1.12-2.13 folds that in GGEC from domestic chickens. The average pepsin potency in GGEC derived from broilers was 15.679 U·mg~(-1), and the corresponding figure for the medicinal from domestic chickens was 26.529 U·mg~(-1). The results of pre-column derivatization reverse-phase high-performance liquid chromatography(RP-HPLC) assay showed that the content of total amino acids and digestion-promoting amino acids in domestic chickens-derived GGEC was 1.12 times and 1.15 times that in GGEC from broilers, and the bitter amino acid content was 1.21 times folds that of the latter. In conclusion, GGEC derived from domestic chickens had weaker fishy odor, stronger enzyme activity, higher content of digestion-promoting amino acids, and stronger bitter taste than GGEC from broilers. This study lays a scientific basis for studying the quality variation of GGEC and provides a method for identifying high-quality GGEC. Therefore, it is of great significance for the development and cultivation of GGEC as both food and medicine and breeding of corresponding varieties.

Biosensor-Coupled In Vivo Mutagenesis and Omics Analysis Reveals Reduced Lysine and Arginine Synthesis To Improve Malonyl-Coenzyme A Flux in Saccharomyces cerevisiae.

Malonyl-coenzyme A (malonyl-CoA) is an important precursor for producing various chemicals, but its low availability limits the synthesis of downstream products in Saccharomyces cerevisiae. Owing to the complexity of metabolism, evolutionary engineering is required for developing strains with improved malonyl-CoA synthesis. Here, using the biosensor we constructed previously, a growth-based screening system that links the availability of malonyl-CoA with cell growth is developed. Coupling this system with in vivo continuous mutagenesis enabled rapid generation of genome-scale mutation library and screening strains with improved malonyl-CoA availability. The mutant strains are analyzed by whole-genome sequencing and transcriptome analysis. The omics analysis revealed that the carbon flux rearrangement to storage carbohydrate and amino acids synthesis affected malonyl-CoA metabolism. Through reverse engineering, new processes especially reduced lysine and arginine synthesis were found to improve malonyl-CoA synthesis. Our study provides a valuable complementary tool to other high-throughput screening method for mutant strains with improved metabolite synthesis and improves our understanding of the metabolic regulation of malonyl-CoA synthesis. IMPORTANCE Malonyl-CoA is a key precursor for the production a variety of value-added chemicals. Although rational engineering has been performed to improve the synthesis of malonyl-CoA in S. cerevisiae, due to the complexity of the metabolism there is a need for evolving strains and analyzing new mechanism to improve malonyl-CoA flux. Here, we developed a growth-based screening system that linked the availability of malonyl-CoA with cell growth and manipulated DNA replication for rapid in vivo mutagenesis. The combination of growth-based screening with in vivo mutagenesis enabled quick evolution of strains with improved malonyl-CoA availability. The whole-genome sequencing, transcriptome analysis of the mutated strains, together with reverse engineering, demonstrated weakening carbon flux to lysine and arginine synthesis and storage carbohydrate can contribute to malonyl-CoA synthesis. Our work provides a guideline in simultaneous strain screening and continuous evolution for improved metabolic intermediates and identified new targets for improving malonyl-CoA downstream product synthesis.