Chemical constituents from the heterotrophic marine microalgae Aurantiochytrium sp. SC145 and their antimicrobial activities.

One new indol, N-methoxymethyltryptophol (1), one new phenolic, (2 R)-2-(4-hydroxyphenyl)ethyl 2-hydroxy-3-phenylpropanoate (2) and fifteen known compounds (3-17) were isolated from the methanol extract of the fermentation of marine microalgae Aurantiochytrium sp. SC145. Their structures were elucidated by 1D-, 2D-NMR spectroscopic analysis, HR-ESI-MS, quantum chemical calculation methods and by comparing their NMR data with those reported in the literature. All compounds were evaluated for their antimicrobial activities against microorganisms. Compounds 2, 3 and 11 significantly exhibited antimicrobial activities on all tested Gram-(+), Gram-(-) bacteria and the yeast C. albicans with MIC values ranging from 32 to 256 µg/mL.

Addressing amino acid-derived inhibitory metabolites and enhancing CHO cell culture performance through DOE-guided media modifications.

Previously, we identified six inhibitory metabolites (IMs) accumulating in Chinese hamster ovary (CHO) cultures using AMBIC 1.0 community reference medium that negatively impacted culture performance. The goal of the current study was to modify the medium to control IM accumulation through design of experiments (DOE). Initial over-supplementation of precursor amino acids (AAs) by 100% to 200% in the culture medium revealed positive correlations between initial AA concentrations and IM levels. A screening design identified 5 AA targets, Lys, Ile, Trp, Leu, Arg, as key contributors to IMs. Response surface design analysis was used to reduce initial AA levels between 13% and 33%, and these were then evaluated in batch and fed-batch cultures. Lowering AAs in basal and feed medium and reducing feed rate from 10% to 5% reduced inhibitory metabolites HICA and NAP by up to 50%, MSA by 30%, and CMP by 15%. These reductions were accompanied by a 13% to 40% improvement in peak viable cell densities and 7% to 50% enhancement in IgG production in batch and fed-batch processes, respectively. This study demonstrates the value of tuning specific AA levels in reference basal and feed media using statistical design methodologies to lower problematic IMs.

Modulation of nutrient precursors for controlling metabolic inhibitors by genome-scale flux balance analysis.

Therapeutic protein productivity and glycosylation pattern highly rely on cell metabolism. Cell culture medium composition and feeding strategy are critical to regulate cell metabolism. In this study, the relationship between toxic metabolic inhibitors and their nutrient precursors was explored to identify the critical medium components toward cell growth and generation of metabolic by-products. Generic CHO metabolic model was tailored and integrated with CHO fed-batch metabolomic data to obtain a cell line- and process-specific model. Flux balance analysis study was conducted on toxic metabolites cytidine monophosphate, guanosine monophosphate and n-acetylputrescine-all of which were previously reported to generate from endogenous cell metabolism-by mapping them to a compartmentalized carbon utilization network. Using this approach, the study projected high level of inhibitory metabolites accumulation when comparing three industrially relevant fed-batch feeding conditions one against another, from which the results were validated via a dose-dependent amino acids spiking study. In the end, a medium optimization design was employed to lower the amount of supplemented nutrients, of which improvements in critical process performance were realized at 40% increase in peak viable cell density (VCD), 15% increase in integral VCD, and 37% increase in growth rate. Tight control of toxic by-products was also achieved, as the study measured decreased inhibitory metabolites accumulation across all conditions. Overall, the study successfully presented a digital twin approach to investigate the intertwined relationship between supplemented medium constituents and downstream toxic metabolites generated through host cell metabolism, further elucidating different control strategies capable of improving cellular phenotypes and regulating toxic inhibitors.

Protein tyrosine phosphatase 1B inhibitors isolated from Morus bombycis.

Bioassay-guided fractionation of the chloroform-soluble fraction of Morus bombycis, using an in vitro PTP1B inhibitory assay led to the identification of three 2-arylbenzofurans, albafuran A (1), mulberrofuran W (2) and mulberrofuran D (6), along with three chalcone-derived Diels-Alder products, kuwanon J (3), kuwanon R (4), and kuwanon V (5). Compounds 1-6 showed remarkable inhibitory activity against PTP1B with IC(50) values ranging from 2.7 to 13.8 microM. Inhibition kinetics were analyzed by Lineweaver-Burk plots, which suggested that compounds 1-6 inhibited PTP1B in a mixed-type manner. The present results indicate that the respective lipophilic and hydroxyl groups of 2-arylbenzofurans and chalcone-derived Diels-Alder products play an important role in inhibition of PTP1B.

Protein tyrosine phosphatase 1B inhibitory by dammaranes from Vietnamese Giao-Co-Lam tea.

ETHNOPHARMACOLOGICAL RELEVANCE: Gynostemma pentaphyllum (Thunb.) tea was used in Vietnamese folk medicine as anti-diabetic agent. AIM OF THE STUDY: This study was aimed to investigate the inhibitory activities of fractions and constituents isolated from Gynostemma pentaphyllum on protein tyrosine phosphatase 1B (PTP1B) since it has been proposed as a treatment therapy for type 2 diabetes and obesity. MATERIALS AND METHODS: The 70% EtOH extract, CHCl3 fraction, EtOAc fraction, BuOH fraction, and seven isolated dammarane triterpenes were evaluated for their inhibitory activity in protein phosphatase enzymes (PTP1B and VHR). RESULTS: CHCl3-soluble fraction showed a dose-dependent inhibitory activity of the PTP1B enzyme with the IC50 value of 30.5 microg/mL. Among seven tested compounds, compounds 6 showed the most potent PTP1B inhibitory activity with IC50 value of 5.3+/-0.4 microM compared to a range 15.7-28.5 microM for the other six compounds. The inhibition mode of 6 was competitive toward p-NPP with a K(i) value of 2.8 microM. CONCLUSION: These study results suggested that the PTP1B inhibitory activity of these dammaranes may enable this plant to play an important role in the treatment of diabetes.

Genistein-derivatives from Tetracera scandens stimulate glucose-uptake in L6 myotubes.

An EtOAc-soluble partition of the MeOH extract of a branch of Tetracera scandens (Dilleniaceae family) was subjected to a glucose-uptake assay, which led to the isolation and identification of five isoflavones of previously known structure namely, genistein (1), its derivatives 3',5'-diprenylgenistein (2), 6,8-diprenylgenistein (3), derrone (4) and alpinumisoflavone (5). Of these, compounds 2--5 exhibited significant glucose-uptake activity in basal and insulin-stimulated L6 myotubes. The findings from adenosine monophosphate-activated kinase (AMPK) activation and glucose transport protein4 (GLUT4) and GLUT1 over-expression revealed certain characteristics of compounds 2--5. These compounds inhibited protein tyrosine phosphatase 1B (PTP1B) activities with IC50 values ranging from 20.63 +/- 0.17 to 37.52 +/- 0.31 microM. No muscle cell toxicity was reported with compounds 3--5, while compounds 1 and 2 reduced muscle cell viability with IC50 values of 34.27 +/- 0.35 and 18.69 +/- 0.19 microM, respectively. It was concluded that T. scandens and its constituents exerted highly desirable activities on type 2 diabetes mellitus treatment since they significantly stimulated the uptake of glucose, AMPK phosphorylation, GLUT4 and GLUT1 mRNA expressions and PTP1B inhibition in L6 myotubes.