Extracellular polysaccharides of endophytic fungus Alternaria tenuissima F1 from Angelica sinensis: Production conditions, purification, and antioxidant properties.

The yield, composition, structure and biofunction of microorganism exopolysaccharide (EPS) depended on their genetics and metabolic pathways. An EPS-producing endophytic fungus was isolated from Angelica sinensis planted in Dingxi, Gansu, China, and identified as Alternaria tenuissima F1. The production optimization, purification and characterization of the purified EPS from A. tenuissima F1 and the in-vitro antioxidant potentials was investigated. Results showed the optimal medium composition were 8% mannose, 2% yeast extract, 0.04% MgSO4·7H2O, 0.005% Vc, and the optimal cultivation conditions were set at initial pH 7.0, culture temperature 30 °C, inoculum size of 3%, 180 r/min for 5 d. The EPS purified by ion exchange column chromatography and gel chromatography was a non-reducing sugar and glycoprotein with pyranoid ring by Fourier transform infrared spectroscopy (FT-IR) analysis, displayed a porous network structure by scanning electron microscope (SEM) observation and exhibited a high thermal stability with the degradation temperature of 303.9 °C by thermogravimetric analysis (TG/DSC). Molecular weight of the purified EPS was 3.246 × 104 g/mol, and EPS was composed of d-galacturonic acid, rhamnose, d-mannose, glucose, and d-galactose in ratio of 0.45:3.02:3.25:1.0:0.95. The EPS exhibited strong scavenging activity and may be a new source of natural antioxidant.

Multi-class mycotoxins analysis in Angelica sinensis by ultra fast liquid chromatography coupled with tandem mass spectrometry.

An ultra fast liquid chromatography coupled with tandem mass spectrometry (UFLC-MS/MS) method was developed and validated for simultaneous analysis of multi-class mycotoxins including aflatoxins (AFB1, AFB2, AFG1 and AFG2), ochratoxin A (OTA), fumonisins (FB1 and FB2) and zearalanone (ZEN) in 20 batches of Angelica sinensis samples collected from different markets and stores in China. The eight mycotoxins were extracted and cleaned up by using QuEChERS-based procedure, and then were quantified under the multiple reaction monitoring (MRM) together with positive and negative ionization modes. Focusing on the optimization of extraction and clean-up conditions, as well as UFLC separation and MS/MS parameters of targeted analytes, the developed method expressed good linearity for the eight mycotoxins within their respective linear ranges with correlation coefficients all higher than 0.9974. The limits of detection (LODs) and quantification (LOQs) ranged from 0.005 to 0.125 µg/kg and from 0.0625 to 0.25 µg/kg, respectively. Recoveries for spiked A. sinensis sample at three different levels were all above 78.9% with relative standard deviations (RSDs) below 6.36% for all analytes. Analysis of real samples demonstrated that two visibly moldy A. sinensis samples were detected with AFB1 of 2.07 and 2.92 µg/kg, and AFG1 of 2.84 and 1.53 µg/kg. The proposed quantitative method with significant advantages including simple pretreatment, rapid determination and high sensitivity would be the preferred candidate for the determination and quantification of multi-class mycotoxin contaminants in complex matrixes, which well fulfilled the maximum residue limits (MRLs) from various countries.

Uncovering potential 'herbal probiotics' in Juzen-taiho-to through the study of associated bacterial populations.

Juzen-taiho-to (JTT) is an immune-boosting formulation of ten medicinal herbs. It is used clinically in East Asia to boost the human immune functions. The active factors in JTT have not been clarified. But, existing evidence suggests that lipopolysaccharide (LPS)-like factors contribute to the activity. To examine this possibility, JTT was subjected to a series of analyses, including high resolution mass spectrometry, which suggested the presence of structural variants of LPS. This finding opened a possibility that JTT contains immune-boosting bacteria. As the first step to characterize the bacteria in JTT, 16S ribosomal RNA sequencing was carried out for Angelica sinensis (dried root), one of the most potent immunostimulatory herbs in JTT. The sequencing revealed a total of 519 bacteria genera in A. sinensis. The most abundant genus was Rahnella, which is widely distributed in water and plants. The abundance of Rahnella appeared to correlate with the immunostimulatory activity of A. sinensis. In conclusion, the current study provided new pieces of evidence supporting the emerging theory of bacterial contribution in immune-boosting herbs.

Terriglobus tenax sp. nov., an exopolysaccharide-producing acidobacterium isolated from rhizosphere soil of a medicinal plant.

An exopolysaccharide-producing bacterium, designated strain DRP 35(T), was isolated from the rhizosphere soil of a medicinal herb, Angelica sinensis, at Geumsan in Korea. Cells were Gram-staining-negative, non-motile, catalase-positive and oxidase-negative short rods. The isolate grew aerobically from 15 to 45 °C (optimum 30 °C), pH 3.5-7.0 (optimum pH 5.0) and in the presence of 0-1.0% (w/v) NaCl. Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain DRP 35(T) belongs to the genus Terriglobus in the phylum Acidobacteria with a sequence similarity of 97.2% and 97.0% to Terriglobus saanensis SP1PR4(T) and Terriglobus roseus KBS63(T), respectively. The genomic DNA G+C content was 62.1 mol%. DNA-DNA relatedness between strain DRP 35(T) and the type strains of the other species of the genus Terriglobus, T. saanensis SP1PR4(T) and T. roseus KBS63(T), were 24.6 and 17.2%, respectively. The predominant menaquinone was MK-8. Major fatty acids were iso-C(15 : 0), C(16 : 1)ω7c and C(16 : 0). The polar lipids were phosphatidylethanolamine, unidentified aminophospholipid and unknown phospholipids. On the basis of polyphasic analysis from this study, strain DRP 35(T) represents a novel species of the genus Terriglobus for which the name Terriglobus tenax sp. nov. is proposed. The type strain is DRP 35(T) ( = KACC 16474(T) = NBRC 109677(T)).

Mucilaginibacter herbaticus sp. nov., isolated from the rhizosphere of the medicinal plant Angelica sinensis.

A strictly aerobic, Gram-staining-negative, non-motile and rod-shaped bacterial strain, DR-9(T), was isolated from rhizosphere soil of the medicinal herb Angelica sinensis. Strain DR-9(T) grew at 20-40 °C, at pH 4.0-9.0 and in the presence of 0-1 % (w/v) NaCl. The major fatty acids were iso-C15 : 0 and summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c and/or iso-C15 : 0 2-OH), MK-7 was the major isoprenoid quinone, and phosphatidylethanolamine and an unidentified aminophospholipid were the major polar lipids. A phylogenetic tree based on 16S rRNA gene sequences showed that strain DR-9(T) formed a lineage within the genus Mucilaginibacter and was closely related to Mucilaginibacter polysacchareus DRP28(T) (96.1 % sequence similarity), Mucilaginibacter myungsuensis HMD1056(T) (95.9 % sequence similarity), Mucilaginibacter ximonensis XM-003(T) (95.8 %) and Mucilaginibacter boryungensis BDR-9(T) (95.1 %). The status of strain DR-9(T) as a representative of a separate species was confirmed by DNA hybridization, with 38.6, 36.3 and 29.9 % DNA-DNA relatedness with M. polysacchareus DRP28(T), M. ximonensis XM-003(T) and M. boryungensis BDR-9(T), respectively. The genomic DNA G+C content of strain DR-9(T) was 49.8 %. These data suggest that strain DR-9(T) should be considered as a representative of a novel species of the genus Mucilaginibacter, for which the name Mucilaginibacter herbaticus sp. nov. is proposed. The type strain is DR-9(T) ( = KACC 16469(T) = NBRC 108839(T)).

[Population structure and ecological distribution of rhizospheric microorganisms of Angelica sinensis].

OBJECTIVE: To study the population structure and ecological distribution of the rhizosperic microorganisms of Angelica sinensis. METHOD: The isolation, culture and identification of microorganisms were studied by the biological methods, and the achieved data were analyzed by the statistical methods for analysis of species diversity. RESULT: With the growing stages of A. sinensis from Min county of Gansu province, Heqing of Yunnan province and Baoxing of Sichuan province, the quantity of rhizosperic microorganisms increased; and it had reduced since October. In the area of Min county Gansu province, the number ratio of bacteria and fungi was higher than that in the other two areas. In addition, the population diversity and dynamic change were different in three areas. In the area of Min county Gansu province, the number of dominant microbial populations and the population diversities of bacteria, actinomyces and fungi in the rhizosphere were greater than those in the other two areas. CONCLUSION: The microecological system and microbial population structure in the rhizosphere of Min county Gansu province were stable. And it was suitable for the growth of A. sinensis in this area.