Determination of Content of 6 Triterpenic Acids in Malusdoumeri Fruit by High-performance Liquid Chromatography

A HPLC method was developed for the simultaneous determination of euscaphic acid, 2α,19α-dihydroxy-3-oxo-urs-12-en-28-oic acid, maslinic acid, corosolic acid, oleanic acid, and ursolic acid in Malus doumeri fruits. Analyses were conducted on the distribution and content of these six triterpenic acids in Malus doumeri fruits from six producing areas in Guangxi. Using gradient elution at a flow rate of 0.8 mL/min, a Diamonsil C18 (2) column was used with a mobile phase of 0.1% formic acid aqueous solution/methanol (v/v). The detection wavelength was 210 nm and the column temperature was 30 ℃. The results showed that the mass concentrations of the six triterpenic acids had a good linear relationship with the chromatographic peak areas in the linear range. The correlation coefficients were greater than 0.9990, the detection limits were within 0.46~2.00 μg/mL, the lower quantitation limits were within 1.53~6.67 μg/mL, the coefficients of variation were less than 5.0%, and the recovery percentages were within 99.2%~101.3%. Six samples from different production areas contained six triterpenic acids (euscaphic acid, 2α,19α-dihydroxy-3-oxo-urs-12-en-28-oic acid, maslinic acid, corosolic acid, oleanic acid, and ursolic acid). Each of the six triterpenic acids was a pentacyclic triterpenic acid. Among the six triterpenic acids, euscaphic acid and 2α,19α-dihydroxy-3-oxo-urs-12-en-28-oic acid were first detected in Malus doumeri fruits. Ursolic acid, euscaphic acid, and 2α,19α-dihydroxy-3-oxo-urs-12-en-28-oic acid were predominant, accounting for 71.82%~81.38% of the total triterpenic acids. The total triterpenic acids content ranged from 808.74 to 1090.75 μg/g FM. The total triterpenic acids content of fruits from Liujiang district of Liuzhou and Pingle of Guilin were relatively high, reaching 1090.75 μg/g FM and 1045.20 μg/g FM, respectively. Overall, this study's HPLC method can detect the triterpenic acids content of Malus doumeri fruits and provide scientific references for the quality evaluation of Malus doumeri fruit's raw materials and products

Minimum Information about a Biosynthetic Gene cluster

A wide variety of enzymatic pathways that produce specialized metabolites in bacteria, fungi and plants are known to be encoded in biosynthetic gene clusters. Information about these clusters, pathways and metabolites is currently dispersed throughout the literature, making it difficult to exploit. To facilitate consistent and systematic deposition and retrieval of data on biosynthetic gene clusters, we propose the Minimum Information about a Biosynthetic Gene cluster (MIBiG) data standard.Chemistry and Chemical Biolog

Identification and utilization of two important transporters: SgvT1 and SgvT2, for griseoviridin and viridogrisein biosynthesis in Streptomyces griseoviridis

Abstract Background Griseoviridin (GV) and viridogrisein (VG, also referred as etamycin), both biosynthesized by a distinct 105 kb biosynthetic gene cluster (BGC) in Streptomyces griseoviridis NRRL 2427, are a pair of synergistic streptogramin antibiotics and very important in treating infections of many multi-drug resistant microorganisms. Three transporter genes, sgvT1–T3 have been discovered within the 105 kb GV/VG BGC, but the function of these efflux transporters have not been identified. Results In the present study, we have identified the different roles of these three transporters, SgvT1, SgvT2 and SgvT3. SgvT1 is a major facilitator superfamily (MFS) transporter whereas SgvT2 appears to serve as the sole ATP-binding cassette (ABC) transporter within the GV/VG BGC. Both proteins are necessary for efficient GV/VG biosynthesis although SgvT1 plays an especially critical role by averting undesired intracellular GV/VG accumulation during biosynthesis. SgvT3 is an alternative MFS-based transporter that appears to serve as a compensatory transporter in GV/VG biosynthesis. We also have identified the γ-butyrolactone (GBL) signaling pathway as a central regulator of sgvT1–T3 expression. Above all, overexpression of sgvT1 and sgvT2 enhances transmembrane transport leading to steady production of GV/VG in titers ≈ 3-fold greater than seen for the wild-type producer and without any notable disturbances to GV/VG biosynthetic gene expression or antibiotic control. Conclusions Our results shows that SgvT1–T2 are essential and useful in GV/VG biosynthesis and our effort highlight a new and effective strategy by which to better exploit streptogramin-based natural products of which GV and VG are prime examples with clinical potential

Enhancement of himastatin bioproduction via inactivation of atypical repressors in Streptomyces hygroscopicus

Three atypical regulatory genes, hmtABD have been discovered within the himastatin biosynthetic gene cluster (BGC) in Streptomyces hygroscopicus ATCC 53653 and the roles of their products have been identified. HmtA and HmtD do not show any structurally distinct features characteristic of regulatory function yet were shown to play important repressive and stimulatory roles, respectively, related to himastatin biosynthesis. HmtB encodes a conserved acetylglutamate kinase; new member of this family serves as repressor of secondary metabolism. Through repressive networks engineering, the limiting functions of HmtA and HmtB along with the activating functions of HmtD in the himastatin BGC have been identified for the first time by gene activation, qPCR, RT-PCR and HPLC studies of selected mutant strains; two of these mutant strains (ΔhmtA and ΔhmtB) produced himastatin in titers (19.02 ± 1.2 μg/mL, 9.9 folds and 30.40 ± 0.83 μg/mL, 15.8 folds) far exceeding those of the wild-type (WT) producer. Overall, this work provides significant insight into secondary metabolic regulatory mechanisms in Streptomyces. These efforts also highlight and validate a new strategy enabling expanded exploitation of cyclopeptidic natural products such as himastatin that demonstrate exciting antimicrobial and antitumor potentials. Keywords: Himastatin, Streptomyces hygroscopicus, Repressor, Biosynthesis, Bioproductio

Identification of the Actinomycin D Biosynthetic Pathway from Marine-Derived <i>Streptomyces costaricanus</i> SCSIO ZS0073

Bioactive secondary metabolites from Streptomycetes are important sources of lead compounds in current drug development. Streptomyces costaricanus SCSIO ZS0073, a mangrove-derived actinomycete, produces actinomycin D, a clinically used therapeutic for Wilm&#8217;s tumor of the kidney, trophoblastic tumors and rhabdomyosarcoma. In this work, we identified the actinomycin biosynthetic gene cluster (BGC) acn by detailed analyses of the S. costaricanus SCSIO ZS0073 genome. This organism produces actinomycin D with a titer of ~69.8 &#956;g mL&#8722;1 along with traces of actinomycin Xo&#946;. The acn cluster localized to a 39.8 kb length region consisting of 25 open reading frames (ORFs), including a set of four genes that drive the construction of the 4-methyl-3-hydroxy-anthranilic acid (4-MHA) precursor and three non-ribosomal peptide synthetases (NRPSs) that generate the 4-MHA pentapeptide semi-lactone, which, upon dimerization, affords final actinomycin D. Furthermore, the acn cluster contains four positive regulatory genes acnWU4RO, which were identified by in vivo gene inactivation studies. Our data provide insights into the genetic characteristics of this new mangrove-derived actinomycin D bioproducer, enabling future metabolic engineering campaigns to improve both titers and the structural diversities possible for actinomycin D and related analogues

Improved Charge Separation and CO₂ Affinity of In₂O₃ by K Doping with Accompanying Oxygen Vacancies for Boosted CO₂ Photoreduction

The CO2 photocatalytic conversion efficiency of the semiconductor photocatalyst is always inhibited by the sluggish charge transfer and undesirable CO2 affinity. In this work, we prepare a series of K-doped In2O3 catalysts with concomitant oxygen vacancies (OV) via a hydrothermal method, followed by a low-temperature sintering treatment. Owing to the synergistic effect of K doping and OV, the charge separation and CO2 affinity of In2O3 are synchronously promoted. Particularly, when P/P0 = 0.010, at room temperature, the CO2 adsorption capacity of the optimal K-doped In2O3 (KIO-3) is 2336 cm3·g–1, reaching about 6000 times higher than that of In2O3 (0.39 cm3·g–1). As a result, in the absence of a cocatalyst or sacrificial agent, KIO-3 exhibits a CO evolution rate of 3.97 μmol·g–1·h–1 in a gas–solid reaction system, which is 7.6 times that of pristine In2O3 (0.52 μmol·g–1·h–1). This study provides a novel approach to the design and development of efficient photocatalysts for CO2 conversion by element doping