Allorhizobium sonneratiae sp. nov., an endophytic bacterium isolated from the root of Sonneratia apetala.

A novel endophytic bacterium, designated strain BGMRC 0089T, was isolated from a surface-sterilized root of Sonneratia apetala. Cells were observed to be Gram-negative, rod-shaped and motile with polar flagella. Strain BGMRC 0089T was found to grow optimally at 28-30 °C, pH 7.0-8.0 and in the presence of 1 % (w/v) NaCl. Strain BGMRC 0089T contained ubiquinone Q-10 and the predominant fatty acid was summed feature 8. The polar lipid profile of strain BGMRC 0089T was found to contain diphosphatidylglycerol, phosphatidylglycerol, phosphatidylmonomethylethanolamine and phosphatidylethanolamine. Based on the results of 16S rRNA gene analysis, this isolate has the closest phylogenetic relationships with Rhizobium lemnae L6-16T (96.5 %) and Allorhizobium oryziradicis N19T (96.4 %). Average nucleotide identity, amino acid identity and digital DNA-DNA hybridization values of the isolate with the type strains of the genera Rhizobium and Allorhizobium were below 84.6, 73.9 and 22.1  %, respectively. Analysis the 4.55 Mb draft genome of strain BGMRC 0089T revealed several plant-associated genes, which may play important roles for the plant in the adaptation to the mangrove habitat. Based on its distinct phylogenetic, phenotypic and chemotaxonomic characteristics, strain BGMRC 0089T is proposed to represent a novel Allorhizobium species, for which the name Allorhizobium sonneratiae sp. nov. is proposed (type strain BGMRC 0089T=DSM 100171T=MCCC 1K04805T).

Novosphingobium album sp. nov., Novosphingobium organovorum sp. nov. and Novosphingobium mangrovi sp. nov. with the organophosphorus pesticides degrading ability isolated from mangrove sediments.

Members of the genus Novosphingobium were frequently isolated from polluted environments and possess great bioremediation potential. Here, three species, designated B2637T, B2580T and B1949T, were isolated from mangrove sediments and might represent novel species in the genus Novosphingobium based on a polyphasic taxonomy study. Phylogenomic analysis revealed that strains B2580T, B1949T and B2637T clustered with Novosphingobium naphthalenivorans NBRC 102051T, 'N. profundi' F72 and N. decolorationis 502str22T, respectively. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between isolates and their closely related species were less than 94 and 54 %, respectively, all below the threshold of species discrimination. The sizes of the genomes of isolates B2580T, B2637T and B1949T ranged from 4.4 to 4.6 Mb, containing 63.3-66.4 % G+C content. Analysis of their genomic sequences identified genes related to pesticide degradation, heavy-metal resistance, nitrogen fixation, antibiotic resistance and sulphur metabolism, revealing the biotechnology potential of these isolates. Except for B2637T, B1949T and B2580T were able to grow in the presence of quinalphos. Results from these polyphasic taxonomic analyses support the affiliation of these strains to three novel species within the genus Novosphingobium, for which we propose the name Novosphingobium album sp. nov. B2580T (=KCTC 72967T=MCCC 1K04555T), Novosphingobium organovorum sp. nov. B1949T (=KCTC 92158T=MCCC 1K03763T) and Novosphingobium mangrovi sp. nov. B2637T (KCTC 72969T=MCCC 1K04460T).

Parvularcula maris sp. nov., an algicidal bacterium isolated from seawater.

A taxonomic study was carried out on strain BGMRC 0090T, which was isolated from seawater. The isolate was a Gram-negative, aerobic, flagellated, rod-shaped bacterium with algicidal activity. Optimal growth was observed at 30 °C, pH 6.0 and with 2 % (w/v) NaCl. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain BGMRC 0090T belonged to the genus Parvularcula, with highest sequence similarity to Parvularcula lutaonensis CC-MMS-1T (98.4 %). Average nucleotide identity, amino acid identity and digital DNA-DNA hybridization values between strain BGMRC 0090T and five strains of the genus Parvularcula with publicly available genomes were below 84.0, 69.2 and 21.4 %, respectively. The genome of strain BGMRC 0090T was 3.2 Mb with 64.8 mol% DNA G+C content and encoded 2905 predicted proteins, three rRNA, 42 tRNA and four ncRNA genes. Some algicidal biosynthesis-associated genes were detected in the genome. Strain BGMRC 0090T contained Q-10 as the major quinone. The predominant fatty acids were identified as summed feature 8 (C18 : 1ω7c/ω6c) and C16 : 0. Based on the polyphasic evidence presented in this paper, strain BGMRC 0090T is concluded to represent a novel species of the genus Parvularcula, for which the name Parvularcula maris sp. nov. is proposed. The type strain is BGMRC 0090T (= KCTC 92591T=MCCC 1K08100T).

Taxonomy and anticancer potential of Streptomyces niphimycinicus sp. nov. against nasopharyngeal carcinoma cells.

Streptomyces species are ubiquitous, Gram-positive, spore-forming bacteria with the ability to produce various clinically relevant compounds. The strain 4503 T was isolated from mangrove sediments, showing morphological and chemical properties which were consistent with those of members of the genus Streptomyces. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the isolate was primarily identified as members of the genus Streptomyces, sharing more than 99% sequence identity to Streptomyces yatensis DSM 41771 T, S. antimycoticus NBRC 12839 T, and S. melanosporofaciens NBRC 13061 T. Average nucleotide identities (ANI) and digital DNA-DNA hybridization (dDDH) values between strain 4503 T and its close relatives were all below 95-96% and 75% of the novel species threshold, respectively. Results from phylogenetic, genomic, phenotypic, and chemotaxonomic characteristics analyses confirmed that the isolate represented a novel species of the genus Streptomyces, for which the name Streptomyces niphimycinicus sp. nov. 4503 T (= MCCC 1K04557T = JCM 34996 T) is proposed. The bioassay-guided fractionation of the extract of strain 4503 T resulted in the isolation of a known compound niphimycin C, which showed cytotoxic activity against nasopharyngeal carcinoma (NPC) cell lines TW03 and 5-8F with half maximal inhibitory concentration (IC50) values of 12.24 µg/mL and 9.44 µg/mL, respectively. Further experiments revealed that niphimycin C not only exhibited the capacity of anti-proliferation, anti-metastasis, induction of cell cycle arrest, and apoptosis, but was also able to increase the reactive oxygen species (ROS) production and regulate several signaling pathways in NPC cells. KEY POINTS: • Strain 4503 T was classified as a novel species of Streptomyces. • Niphimycin C correlates with the cytotoxic effect of strain 4503 T against NPC cells. • Niphimycin C induces apoptosis, autophagic flux disruption and cell cycle arrest.

Novosphingobium beihaiensis sp. nov., a novel pesticide-tolerant bacterium isolated from mangrove sediments.

A novel Novosphingobium species, designated strain B2638T, was isolated from mangrove sediments which was collected from Beibu Gulf, Beihai, P. R. China. The isolate could grow in the presence of chlorpyrifos. Phylogenetic analysis based on 16S rRNA gene sequence revealed that the isolate belonged to the genus Novosphingobium, showing 99.9% sequence similarity with N. decloroationis 502str22T and less than 98% similarity with other type strain of species of this genus. Molecular typing by BOX-PCR divided strain B2638T and N. declorationis 502str22T into two clusters and indicated that they were not identical. Genomic comparison referenced by values of the DNA-DNA hybridization (dDDH) and the average nucleotide identity (ANI) between strain B2638T and its close phylogenetic neighbors were 20.0-29.5% and 75.3-85.3%, respectively, that were lower than proposed thresholds for bacterial species delineation. The major fatty acids (> 10%) were identified as C18:1 ω7c, C17:1 iso ω9c and C16:0. The main polar lipids contained diphosphatidylglycerol, phosphatidylethanolamine, sphingoglycolipid, phosphatidyl glycerol, unidentified lipid and unidentified aminolipid. Results from phenotypic, chemotaxonomic and genotypic analyses proposed that strain B2638T (= MCCC 1K07406T = KCTC 72968 T) is represented a novel species in the genus Novosphingobium, for which the names Novosphingobium beihaiensis sp. nov. is proposed.

Mycolicibacterium aurantiacum sp. nov. and Mycolicibacterium xanthum sp. nov., two novel actinobacteria isolated from mangrove sediments.

Two novel actinobacteria with the ability to degrade kerosene, designated as B3033T and Y57T, were isolated from mangrove sediments in Tieshan Harbour, South China Sea. Both strains are Gram-staining-positive, non-spore forming, slow-growing, oxidase-positive, non-motile and aerobic. Their major cellular fatty acids were C16 : 0 and C18 : 1ω9c. Analysis of 16S rRNA gene sequences revealed the close relationship of strain B3033T to Mycobacterium kyogaense DSM 107316T (99.4 % nucleotide identity) and strain Y57T to Mycolicibacterium chubuense ATCC 27278T (98.7 %) and Mycolicibacterium rufum JS14T (98.7 %). Whole genome average nucleotide blast identity (ANI) and the digital DNA-DNA hybridization (dDDH) values between the two isolates and the type strains of species of the genus Mycolicibacterium were lower than 94 and 45 %, respectively, which were below the threshold values of 95 % (for ANI) and 70 % (for dDDH) recommended for bacterial species differentiation. The genome sequence of B3033T comprised a circular 11.0 Mb chromosome with a DNA G+C content of 68.1 mol%. Y57T had a genome size of 5.6 Mb and a DNA G+C content of 68.7 mol%. Genes involved in degradation of aromatic compounds and copper resistance were identified in the genomes of both strains that could improve their adaptive capacity to the mangrove environment. These results combined with those of chemotaxonomic analyses, MALDI-TOF MS profiles and phenotypic analyses support the affiliation of these strains to two novel species within the genus Mycolicibacterium, for which we propose the names Mycolicibacterium aurantiacum sp. nov. B3033T (=KCTC 49712T=MCCC 1K04526T) and Mycolicibacterium xanthum sp. nov. Y57T (=KCTC 49711T=MCCC 1K04875T) as type strains.

Roseomonas coralli sp. nov., a heavy metal resistant bacterium isolated from coral.

A Gram-stain-negative, aerobic, mesophilic, non-motile bacterium, designated M0104T, was isolated from a gorgonian coral collected from Xieyang island, Guangxi Province, PR China. Colonies of the strain were non-motile cocci and pink. The strain grew at 15-34 °C (optimum, 28 °C), pH 4.5-8.0 (optimum, pH 7.0) and with 0-4% (w/v) NaCl (optimum, 0-2 %). Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain M0104T was closely related to Roseomonas deserti JCM 31275T (96.2 %), Roseomonas vastitatis KCTC 62043T (96.0 %), Roseomonas aerofrigidensis JCM 31878T (95.9 %) and Roseomonas oryzae KCTC 42542T (95.7 %). The strain had an assembly size of 5.0 Mb and a G+C content of 71.0mol%. Genes involved in copper, cadmium, lead, arsenic and zinc resistance were identified in the genome of strain M0104T. The digital DNA-DNA hybridization and average nucleotide identity values between the genome sequence of strain M0104T and those of closely related type strains were 19.4-24.9 % and 74.3-81.8 %, respectively. Strain M0104T contained C18:1 ω7c, C18:3 ω3c, anteiso C11:0 and C16:0 as the major fatty acids (>7 %) and ubiquinone Q-10 as the sole isoprenoid quinone. Diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and phosphatidylcholine were its major polar lipids. Based on its phenotypic, phylogenetic and chemotaxonomic properties, strain M0104T is proposed to represent a novel species within the genus Roseomonas, for which the name Roseomonas coralli sp. nov. is proposed. The type strain is M0104T (=KCTC 62359T=MCCC 1K03632T).

Acuticoccus mangrovi sp. nov., with an antibacterial property, isolated from mangrove sediment.

A Gram-stain-negative, aerobic, milky white bacterium, designated B2012T, was isolated from mangrove sediment collected at Beibu Gulf, South China Sea. Antimicrobial activity assay revealed that the isolate possesses the capability of producing antibacterial compounds. Strain B2012T shared the highest 16S rRNA gene sequence relatedness (96.9-95.5 %) with members of the genus Acuticoccus. The isolate and all known Acuticoccus species contain Q-10 as the main respiratory quinone and have the same polar lipid components (phosphatidylcholine, unidentified glycolipid, unidentified lipid, unidentified amino lipid and phosphatidylglycerol). However, genomic relatedness referred by values of average nucleotide identity, digital DNA-DNA hybridization, average amino acid identity and the percentage of conserved proteins between strain B2012T and other type strains of the genus Acuticoccus were below the proposed thresholds for species discrimination. The genome of strain B2012T was assembled into 65 scaffolds with an N50 size of 244239 bp, resulting in a 5.5 Mb genome size. Eight secondary metabolite biosynthetic gene clusters were detected in this genome, including three non-ribosomal peptide biosynthetic loci encoding yet unknown natural products. Strain B2012T displayed moderately halophilic and alkaliphilic properties, growing optimally at 2-3 % (w/v) NaCl concentration and at pH 8-9. The major cellular fatty acids (>10 %) were anteiso-C15 : 0, C16 : 0 dimethyl aldehyde (DMA) and C16 : 0. Combined data from phenotypic, genotypic and chemotaxonomic analyses suggested that strain B2012T represents a novel species of the genus Acuticoccus, for which the name Acuticoccus mangrovi sp. nov. is proposed. The type strain of the type species is B2012T (=MCCC 1K04418T=KCTC 72962T).

Mesobaculum littorinae gen. nov., sp. nov., a novel bacterium isolated from a sea snail Littorina scabra.

Members within the family Rhodbacteraceae are morphologically and genetically highly diverse, and originate mostly from coastal marine environments. In this study, a novel species of this family, designated M0103T, was isolated from the surface of a sea snail Littorina scabra. Strain M0103T is Gram-stain-negative, halophilic, non-motile and non-Bacteriochlorophyll a-producing bacterium. Several phenotypic characteristics of the isolate were similar to other species within this family, such as the sole respiratory quinone Q-10 and major fatty acid components C18 : 1 ω7c, C18 : 0 and C16 : 0. Strain M0103T contains a diphosphatidylglycerol, a phosphatidylglycerol, a phosphatidylcholine, a phosphatidy ethanolamine, a phosphatidylinositol, five unidentified phospholipids and four unidentified polar lipids. Based on the 16S rRNA gene sequence analysis, this isolate showed the closest phylogenetic relationship with 'Palleronia pontilimi' GH1-23T (95.1 %). Values of average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) of genome sequences were of 70.1-76.4 % and 18.3-20.9 % between the isolate and 24 closely related type strains. Analysis the 4.0 Mb genome of strain M0103T revealed several putative genes associated with cellular stress resistance, which may play protective roles for the isolate in the adaptation to a marine environment. Phylogenetic, phenotypic and chemotaxonomic analyses suggested that strain M0103T represents a novel genus and novel species of the family Rhodobacteraceae, for which the name Mesobaculum littorinae gen. nov., sp. nov. is proposed. The type strain is M0103T (=MCCC 1K03619T=KCTC 62358T).

Thermohalobaculum xanthum gen. nov., sp. nov., a moderately thermophilic bacterium isolated from mangrove sediment.

A novel moderately thermophilic and halophilic bacterium, designated strain M0105T, was isolated from mangrove sediment collected in the Beibu Gulf, south China. The isolate is Gram-negative, non-motile and rod-shaped bacterium with smooth colonies of pale-yellow appearance. Growth occurs at 15-46 °C (optimum 37-40 °C) and pH range of 6.0-10.0 (optimum pH 8.0-9.0). It required 1-7% NaCl (optimum 3-5%) for growth. Strain M0105T was affiliated to the family 'Rhodobacteraceae', sharing the highest 16S rRNA gene sequence similarity with Limibaculum halophilum CAU 1123T (96.8%). The major menaquinone Q-10 and the dominant unsaturated fatty acid (C18:1ω7) in this family were also detected in the strain M0105T. The genome sequence possesses a circular 4.1 Mb chromosome with a G + C content of 67.9%. Strain M0105T encoded many genes for cellular stress resistance and nutrient utilization, which could improve its adaptive capacity to the mangrove environment. Values of conserved proteins (POCP), average nucleotide identity, average amino acid identity (AAI) and DNA-DNA hybridization (dDDH) between the isolate and closely related species were below the proposed threshold for species discrimination. Information from phenotypic, chemotaxonomic and phylogenetic analyses proposed that strain M0105T should be assigned to a novel genus within the family 'Rhodobacteraceae'. Thus, we suggested that the strain M0105T represents a novel species in a new genus, for which the name Thermohalobaculum xanthum gen. nov., sp. nov. is proposed. The type strain of the type species is M0105T (= BGMRC 2019T = KCTC 52118T = MCCC 1K03767T = NBRC 112057T).

Novel Marine Secondary Metabolites Worthy of Development as Anticancer Agents: A Review.

Secondary metabolites from marine sources have a wide range of biological activity. Marine natural products are promising candidates for lead pharmacological compounds to treat diseases that plague humans, including cancer. Cancer is a life-threatening disorder that has been difficult to overcome. It is a long-term illness that affects both young and old people. In recent years, significant attempts have been made to identify new anticancer drugs, as the existing drugs have been useless due to resistance of the malignant cells. Natural products derived from marine sources have been tested for their anticancer activity using a variety of cancer cell lines derived from humans and other sources, some of which have already been approved for clinical use, while some others are still being tested. These compounds can assault cancer cells via a variety of mechanisms, but certain cancer cells are resistant to them. As a result, the goal of this review was to look into the anticancer potential of marine natural products or their derivatives that were isolated from January 2019 to March 2020, in cancer cell lines, with a focus on the class and type of isolated compounds, source and location of isolation, cancer cell line type, and potency (IC50 values) of the isolated compounds that could be a guide for drug development.

Parallel micro-Raman spectroscopy of multiple cells in a single acquisition using hierarchical sparsity.

Parallel micro-Raman spectroscopy can significantly expand the analytical capacity of single biological cells. By positioning the Raman spectra of multiple trapped cells on a detector array along the grating dispersion direction, the throughput of single-cell analysis can be improved by orders of magnitude. However, accurate retrieval of the individual spectra from the superimposed spectrum in a single acquisition presents great challenges. In this work, we developed a hierarchical sparsity method under a compressive sensing framework. The method combined a group-selection strategy with in-group sparsity for spectral reconstruction. The performances of the developed method were demonstrated with both simulated and experimental data, and the Raman spectra of the individual trapped cells were retrieved with both high accuracy and low noises; especially, with a group-selection mechanism, the developed method successfully avoided wrong selection of the eigenspectra for spectral reconstruction. The technique is expected to find wide applications in simultaneous monitoring of long biological processes of multiple cells by Raman spectroscopy.

Antibiotics Development and the Potentials of Marine-Derived Compounds to Stem the Tide of Multidrug-Resistant Pathogenic Bacteria, Fungi, and Protozoa.

As the search for new antibiotics continues, the resistance to known antimicrobial compounds continues to increase. Many researchers around the world, in response to antibiotics resistance, have continued to search for new antimicrobial compounds in different ecological niches such as the marine environment. Marine habitats are one of the known and promising sources for bioactive compounds with antimicrobial potentials against currently drug-resistant strains of pathogenic microorganisms. For more than a decade, numerous antimicrobial compounds have been discovered from marine environments, with many more antimicrobials still being discovered every year. So far, only very few compounds are in preclinical and clinical trials. Research in marine natural products has resulted in the isolation and identification of numerous diverse and novel chemical compounds with potency against even drug-resistant pathogens. Some of these compounds, which mainly came from marine bacteria and fungi, have been classified into alkaloids, lactones, phenols, quinones, tannins, terpenes, glycosides, halogenated, polyketides, xanthones, macrocycles, peptides, and fatty acids. All these are geared towards discovering and isolating unique compounds with therapeutic potential, especially against multidrug-resistant pathogenic microorganisms. In this review, we tried to summarize published articles from 2015 to 2019 on antimicrobial compounds isolated from marine sources, including some of their chemical structures and tests performed against drug-resistant pathogens.

A novel MyD88 inhibitor LM9 prevents atherosclerosis by regulating inflammatory responses and oxidative stress in macrophages.

Development of atherosclerosis involves chronic and sustained inflammation and oxidative stress. Recent studies have linked atherosclerosis to the innate immune system. Genetic deficiency in myeloid differentiation primary-response protein 88 (MyD88) protects against the development and progression of atherosclerosis. However, it is unknown if pharmacological inhibition of MyD88 is able to be a therapeutic strategy for this disease. In this study, we evaluated the effect of a newly synthesized small-molecule inhibitor of MyD88, LM9, in an ApoE-/- mouse model of atherosclerosis. Our results showed that the major source of MyD88 in atherosclerotic lesions is infiltrated macrophage. Treatment of HFD-fed ApoE-/- mice with LM9 significantly attenuated the pathogenesis of atherosclerosis, accompanied with reduced vascular inflammatory responses and oxidative stress. These effects were achieved without changes to serum lipid levels. We further showed that LM9 inhibited oxidized-lipoprotein induced foam cell formation through suppression of MyD88 and inflammatory pathway in macrophages. Additionally, either LM9 treatment or MyD88 knockdown prevented ox-LDL-induced oxidative stress in macrophages. This study highlights the translational role of MyD88 as a therapeutic target and identifies the MyD88 inhibitor LM9 as a new candidate for the treatment of atherosclerosis.

Characterization of a bifunctional alginate lyase as a new member of the polysaccharide lyase family 17 from a marine strain BP-2.

OBJECTIVES: Bifunctional alginate lyase can efficiently saccharify alginate biomass and prepare functional oligosaccharides of alginate. RESULTS: A new BP-2 strain that produces alginate lyase was screened and identified from rotted Sargassum. A new alginate lyase, Alg17B, belonging to the polysaccharide lyase family 17, was isolated and purified from BP-2 fermentation broth by freeze-drying, dialysis, and ion exchange chromatography. The enzymatic properties of the purified lyase were investigated. The molecular weight of Alg17B was approximately 77 kDa, its optimum reaction temperature was 40-45 °C, and its optimum reaction pH was 7.5-8.0. The enzyme was relatively stable at pH 7.0-8.0, with a temperature range of 25-35 °C, and the specific activity of the purified enzyme reached 4036 U/mg. A low Na+ concentration stimulated Alg17B enzyme activity, but Ca2+, Zn2+, and other metal ions inhibited it. Substrate specificity analysis, thin-layer chromatography, and mass spectrometry showed that Alg17B is an alginate lyase that catalyses the hydrolysis of sodium alginate, polymannuronic acid (polyM) and polyguluronic acid to produce monosaccharides and low molecular weight oligosaccharides. Alg17B is also bifunctional, exhibiting both endolytic and exolytic activities toward alginate, and has a wide substrate utilization range with a preference for polyM. CONCLUSIONS: Alg17B can be used to saccharify the main carbohydrate, alginate, in the ethanolic production of brown algae fuel as well as in preparing and researching oligosaccharides.

AlgM4: A New Salt-Activated Alginate Lyase of the PL7 Family with Endolytic Activity.

Alginate lyases are a group of enzymes that catalyze the depolymerization of alginates into oligosaccharides or monosaccharides. These enzymes have been widely used for a variety of purposes, such as producing bioactive oligosaccharides, controlling the rheological properties of polysaccharides, and performing structural analyses of polysaccharides. The algM4 gene of the marine bacterium Vibrio weizhoudaoensis M0101 encodes an alginate lyase that belongs to the polysaccharide lyase family 7 (PL7). In this study, the kinetic constants Vmax (maximum reaction rate) and Km (Michaelis constant) of AlgM4 activity were determined as 2.75 nmol/s and 2.72 mg/mL, respectively. The optimum temperature for AlgM4 activity was 30 °C, and at 70 °C, AlgM4 activity dropped to 11% of the maximum observed activity. The optimum pH for AlgM4 activity was 8.5, and AlgM4 was completely inactive at pH 11. The addition of 1 mol/L NaCl resulted in a more than sevenfold increase in the relative activity of AlgM4. The secondary structure of AlgM4 was altered in the presence of NaCl, which caused the α-helical content to decrease from 12.4 to 10.8% and the ß-sheet content to decrease by 1.7%. In addition, NaCl enhanced the thermal stability of AlgM4 and increased the midpoint of thermal denaturation (Tm) by 4.9 °C. AlgM4 exhibited an ability to degrade sodium alginate, poly-mannuronic acid (polyM), and poly-guluronic acid (polyG), resulting in the production of oligosaccharides with a degree of polymerization (DP) of 2-9. AlgM4 possessed broader substrate, indicating that it is a bifunctional alginate lyase. Thus, AlgM4 is a novel salt-activated and bifunctional alginate lyase of the PL7 family with endolytic activity.

Spectrum-effect relationship between HPLC fingerprints and hypolipidemic effect of Curcuma aromatica.

Curcuma aromatica is used as a traditional Chinese medicine, and it is mainly distributed in Guangxi, China. In this study, 10 batches of C. aromatica were collected from different origins in Guangxi. The fingerprints were established by HPLC technique to investigate the quality stability of C. aromatica. The spectrum-effect relationship between HPLC fingerprints and hypolipidemic effect of C. aromatica was assessed by similarity analysis, gray relational analysis and multiple linear regression analysis. From the results, the similarity values between each batch of C. aromatica and reference fingerprint were >0.880, indicating the good quality stability of the 10 batches of C. aromatica. Twenty common peaks were selected as the fingerprints to evaluate the quality and hypolipidemic effect of C. aromatica. The results of spectrum-effect relationship showed that peaks 10, 18, 13, 15 and 17 in the fingerprints were closely related to hypolipidemic effect. This study successfully established the spectrum-effect relationship between HPLC fingerprints and hypolipidemic effect of C. aromatica, which provided methods for quality control and more effectively studies on bioactive compounds of C. aromatica. It could also provide a new simple and effective method for utilizing the fingerprints to optimize the Chinese prescription and develop traditional Chinese medicine.

[Apoptosis of hepatocellular induced by active ingredients of Zanthoxyli Radix by Raman spectroscopy].

The apoptosis of mono-hepatocellular induced by the active ingredients of the Zanthoxyli Radix was investigated using laser Raman spectroscopy. Hepatoma cells (BEL-7404) were treated with 10 mg•L⁻¹ nitidine chloride and 3 g•L⁻¹ the extracts of Zanthoxyli Radix, respectively, then were divided into two parts, one for fluorescence staining, the other for determination of Raman spectroscopy. The acquired spectra were then processed by background elimination, smoothing, and normalization. Fluorescence staining results showed that the nucleuses from untreated group were uniformly stained, while those from the group treated for 48 hours were densely stained and broken. The spectra results revealed that the intensity of peaks associated with nucleic acid and protein decreased after the cells were incubated with the extracts of Zanthoxyli Radix for 12, 24, 36 and 48 hours. The intensity of peaks at 785,1 002,1 175,1 660 cm⁻¹ was decreased with the time of the cells were incubated by the extracts of Zanthoxyli Radix. The results indicated that the extracts of Zanthoxyli Radix could induce the apoptosis of hepatoma cells and reduce the amount of nucleic acid and protein in the cells. There is a certain relevance between the drug treatment time and the efficacy. The above results suggest that Raman spectra can provide abundant information about the changes in biological macromolecules within the cells after incubated by the extracts of Zanthoxyli Radix and serve as an effective method for the real time measurement of apoptosis.

[A Modified Procedure to Isolate Synchronous Cells from Yeasts with Continuous Percoll Density Gradient and Their Raman Discrimination].

A modified procedure of Percoll density gradient centrifugation was developed to isolate and fractionate synchronous cells from stationary phase (sp) cultures of different yeast strains, as well as Raman spectra discrimination of single yeast cells was reported. About 1.75 mL Percoll solution in 2 mL polypropylene centrifugal tube was centrifuged at 19,320 g, 20 °C with an angle rotor for 15 min to form continuous densities gradient (1.00~1.31 g · mL(-1)), approximately 100 µL sample was overlaid onto the preformed continuous density gradient carefully, subsequently, centrifuged at 400 g for 60 min in a tabletop centrifuge equipped with a angle rotor at 25 °C. Yeast samples could be observed that the suspensions were separated into two cell fractions obviously. Both fractions of different yeast strains were respectively determined by differential interference contrast (DIC), phase contrast microscope and synchronous culture to distinguish their morphological and growth trait. The results showed that the lower fraction cells were unbudded, mostly unicellular, highly refractive, homogeneous and uniform in size, and represented growth characteristic synchronously; Their protoplasm had relatively high density, and contained significant concentrations of glycogen; all of which were accordant with description of quiescent yeast cells and G0 cells in previously published paper. It was shown that lower fraction was quiescent cells, synchronous G0 cells as well. A Raman tweezers setup was used to investigate the differences between two fractions, G0 cells and non G0 cells, at a single cell level. The result showed that both G0 cells and the non G0 cells had the same characteristic peaks corresponding biological macromolecules including proteins, carbohydrates and nucleic acids, but all characteristic peak intensities of G0 cells were higher than that of non G0 cells, implied that the macromolecular substance content of G0 cells was more higher. Principal component analysis (PCA) was performed between G0 cells and non G0 cells, the results showed that the chemical composition content among the synchronization G0 cells has less difference, and G0 cells were homogeneous but non G0 cells were heterogeneous, indicating single cell optical tweezers Raman spectroscopy could identify the synchronous and asynchronous cells. The modified method is feasible, economical and efficient highly. G0 synchronous cells of most yeast strains could be isolated by a modification of Percoll density gradient centrifugation.

[Raman spectra of endospores of Bacillus subtilis by alkali stress].

To research the lethal mechanism of spores stressed by alkali, laser tweezers Raman spectroscopy (LTRS) combined with principal components analysis (PCA) was used to study the physiological process of single spore with alkali stress. The results showed that both spores and germinated spores had tolerance with alkali in a certain range, but the ability of spores was obviously lower than that of spores due to the release of their Ca2+ -DPA which plays a key role in spores resistance as well as spores resistance to many stresses; A small amount of Ca2+ -DPA of spores was observed to release after alkali stress, however, the behavior of release was different with the normal Ca2+ -DPA release behavior induced by L-alanine; The data before and after alkali stress of the spores and g. spores with PCA reflected that alkali mainly injured the membrane of spores, and alkali could be easily enter into the inner structure of spores to damage the structure of protein backbone and injure the nucleic acid of spores. We show that the alkali could result in the small amount of Ca2+ -DPA released by destroying the member channel of spores.