Evaluation of microalgae and cyanobacteria as potential sources of antimicrobial compounds

In recent decades, marine microorganisms have become known for their ability to produce a wide variety of secondary bioactive metabolites. Several compounds have been isolated from marine microorganisms for the development of novel bioactives for the food and pharmaceutical industries. In this study, a number of microalgae were evaluated for their antimicrobial activity against gram-positive and gram-negative bacteria, including food and plant pathogens, using various extraction techniques and antimicrobial assays. Disc diffusion and spot-on-lawn assays were conducted to confirm the antimicrobial activity. To measure the potency of the extracts, minimum inhibition concentrations (MIultCs) were measured. Three microalgae, namely Isochrysis galbana, Scenedesmus sp. NT8c, and Chlorella sp. FN1, showed strong inhibitory activity preferentially against gram-positive bacteria. These microalgal species were then selected for further purification and analysis, leading to compound identification. By using a mixture of different chromatography techniques gas chromatography–mass spectrometry (GC–MS) and high-performance liquid chromatography (HPLC) and ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS), we were able to separate and identify the dominant compounds that are responsible for the inhibitory activity. Additionally, nuclear magnetic resonance (NMR) was used to confirm the presence of these compounds. The dominant compounds that were identified and purified in the extracts are linoleic acid, oleic acid, docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA). These compounds are the potential candidates that inhibit the growth of gram-positive bacteria. This indicates the potential use of microalgae and their antimicrobial compounds as biocontrol agents against food and plant pathogens.     

Efficient identification of fungal antimicrobial principles by tandem MS and NMR database

The continuous re-isolation of the known and non-applicable compounds that is time-consuming and wasting resources is still a critical problem in the discovery of bioactive entities from natural resources. To efficiently address the problem, high performance liquid chromatography-diode array detector-microfractionation (HPLC-DAD-microfractionation) guided by disk agar diffusion assay was developed, and the active compounds were further identified using the tandem mass spectrometry (MS/MS)-based molecular networking. Of 150 fungal strains screened, the methanolic extracts of Phoma herbarum PPM7487, Cryptosporiopsis ericae PPM7405, and Albifimbria verrucaria PPM945 exhibited potent antimicrobial activity against Candida albicans SC5314 and Cryptococcus neoformans H99 in the preliminary agar diffusion assay. The concept of OSMAC (one strain many compounds) was employed in the fungal cultures in order to enrich the diversity of the 2^nd metabolites in this study. HPLC coupled with off-line bioactivity-directed profiling of the extracts enabled a precise localization of the compounds responsible for the conspicuous antimicrobial activity. The purified active compounds were identified based mainly on MS/MS database, and further supported by ¹³C nuclear magnetic resonance (NMR) spectral data compared to the literatures. In addition to nineteen known compounds, a new trichothecene derivative 1, namely trichoverrin D, was isolated and identified through this protocol. The antifungal activities of all the pure isolates were evaluated, and the structure activity relationships were also inferred. This report has demonstrated the combination of HPLC microfractination and MS/MS coupled by NMR spectral dereplication for speeding up the antimicrobial natural products discovery process.     

草血竭低极性部位化学成分的气相色谱-质谱法分析

目的 研究草血竭(Polygonum paleaceum Wall)低极性部位的主要化学成分。 方法 草血竭用乙醇提取,提取液回收乙醇后用石油醚萃取,采用气相色谱-质谱法(GC-MS)对草血竭低极性部位进行分析测定,与标准谱库进行比较分析,并用峰面积归一化法计算各成分的质量百分数。 结果 共检出40个化学成分,其中主要成分β-瑟林烯和反油酸乙酯含量较高。 结论 草血竭低极性部位主要成分为β-瑟林烯(59.32%)和反油酸乙酯(27.02%),并含有棕榈酸(4.18%)、13,27-cycloursan-3-one(2.55%)、顺式-11-二十碳烯酸(1.31%)等化学成分。     

The Electrospray Ionization - Mass Spectra of Erythromycin A Obtained from a Marine Streptomyces sp. Mutant

In our ongoing search for production improvements of bioactive secondary metabolites from marine Streptomyces through the induction of mutations using UV light, out of 145 isolates, mutant 10/14 was able to produce potent antibacterial metabolites other than the parent strain as established by chromatographic analysis. Up-scaling fermentation of mutant 10/14, followed by working up and isolation delivered five metabolites, phenazine, 1-acetyl-β -carboline, perlolyrin and erythromycin A, along with an oily substance. The latter two compounds were responsible for the antibacterial activity of the strain. In this article, we discuss with the mutation of the marine Streptomyces sp. AH2, bioactivity evaluation, fermentation and isolation of the microbial metabolites. Moreover, we study to first time in detail the 1D and 2D NMR and ESI MS data including ESI MS² and MS³ patterns combined with HRESI MS of erythromycin A.     

哈士蟆卵油化学成分研究(Ⅱ)

目的分离鉴定哈士蟆卵油化学成分。方法硅胶、LH 2 0柱色谱分离 ,通过IR、NMR及MS光谱分析法确定结构。结果分得 6个化合物 ,分别鉴定为 (Ⅰ )棕榈酸 α 单甘油酯、(Ⅱ )棕榈酸 α,α′ 甘油二酯、(Ⅲ )油酸 α 单甘油酯、(Ⅳ )硬脂酸、(Ⅴ )棕榈酰胺和 (Ⅵ )棕榈酸乙酯及硬脂酸乙酯。结论 6个化合物均为首次从哈士蟆卵中分离得到。     

Synthesis of platensimycin analogues and their antibiotic potency

Platensimycin is a natural product isolated from various strains of Streptomyces platensis which exhibits antimicrobial activity against Gram positive bacteria, including vancomycin- and linezolide-resistant species. Analogues of platensimycin were synthesized from 3-aminobenzoic acid or other aniline derivatives and several alkyl- and aryl-carboxylic acids. The resulting compounds were tested in an agar diffusion assay against several bacteria and fungi.     

[1,2,4]Triazolo[4,3-a]quinoxaline: synthesis, antiviral, and antimicrobial activities

A series of novel [1,2,4]triazolo[4,3-a]quinoxaline derivatives and their isosteres, pyrimido-quinoxaline, were synthesized as potential antiviral and antimicrobial agents. The new compounds were synthesized via aromatic nucleophilic substitution of 4-chloro-8-methyl[1,2,4]triazolo[4,3-a]quinoxaline-1-amine with different amines and triazole-2-thiol. Some of the synthesized compounds were subjected to antiviral and cytotoxicity screening using plaque-reduction assay. Most of the tested compounds exhibited cytotoxicity at concentration 160?μg/ml and compound 8b showed promising antiviral activity. In vitro antimicrobial screening against different pathogenic organisms using agar diffusion method demonstrated that compounds 4d, 6c, 7b, and 8a exhibit antibacterial and/or antifungal activities.     

Antityrosinase and antimicrobial activities from Thai medicinal plants

Various dermatological disorders and microbial skin infection can cause hyperpigmentation. Therefore, screenings for whitening and antimicrobial agents from Thai medicinal plants have been of research interest. Seventy-seven ethanol plant extracts were investigated for antityrosinase activity, eleven samples showed the tyrosinase inhibition more than 50 % were further preliminary screening for antimicrobial activity by agar disc diffusion and broth micro-dilution methods. Artocarpus integer (Thunb.) Merr. (Moraceae) root extract, which showed the potential of tyrosinase inhibition with 90.57 ± 2.93 % and antimicrobial activity against Staphylococcus aureus, S. epidermidis, Propionibacterium acnes and Trichophyton mentagophytes with inhibition zone as 9.10 ± 0.00, 10.67 ± 0.09, 15.25 ± 0.05 and 6.60 ± 0.17 mm, respectively was selected for phytochemical investigation. Three pure compounds were isolated as artocarpin, cudraflavone C and artocarpanone. And artocarpanone exhibited anti-tyrosinase effect; artocarpin and cudraflavone C also showed the potential of antibacterial activity against S. aureus, S. epidermidis and P. acnes with MIC at 2, 4 and 2 μg/ml, respectively and MBC at 32 μg/ml for these bacteria. So, these pure compounds are interesting for further study in order to provide possibilities of new whitening and antibacterial development. This will be the first report of phytochemical investigation of A. integer root.     

海洋链霉菌Streptomyces sp. S598中抗菌活性产物的研究

目的 对海洋链霉菌Streptomyces sp. S598固体发酵提取物的抗菌成分进行研究。方法 采用葡聚糖凝胶Sephadex LH-20、反相柱色谱、硅胶柱层析及半制备HPLC等方法分离纯化该菌株的活性次级代谢产物,并通过ESI-MS、NMR等波谱技术结合文献对比鉴定其结构。结果 从10L固体发酵提取物中分离得到5个单体化合物,分别鉴定为缬氨霉素(1)、二活菌素(2)、亚油酸(3)、亚油酸甲酯(4)和油酸(5)。结论 海洋链霉菌Streptomyces sp. S598能代谢产生具有多种生物活性的化合物。化合物1对MRSA 28300和白念珠菌(ATCC10231)的MIC为1μg/mL,化合物2具有较强的广谱抗细菌活性,对肺炎链球菌、流感嗜血菌的MIC为0.125μg/mL,尤其具有显著的抗MRSA的活性,对MRSA 28300的MIC<0.125μg/mL。化合物1和2均为首次从该种放线菌中分离得到,且本文首次报道其对MRSA均具有抑制活性。     

海洋链霉菌Streptomyces sp. S598中抗菌活性产物的研究

目的 对海洋链霉菌Streptomyces sp. S598固体发酵提取物的抗菌成分进行研究。方法 采用葡聚糖凝胶Sephadex LH-20、反相柱色谱、硅胶柱层析及半制备HPLC等方法分离纯化该菌株的活性次级代谢产物,并通过ESI-MS、NMR等波谱技术结合文献对比鉴定其结构。结果 从10L固体发酵提取物中分离得到5个单体化合物,分别鉴定为缬氨霉素(1)、二活菌素(2)、亚油酸(3)、亚油酸甲酯(4)和油酸(5)。结论 海洋链霉菌Streptomyces sp. S598能代谢产生具有多种生物活性的化合物。化合物1对MRSA 28300和白念珠菌(ATCC10231)的MIC为1μg/mL,化合物2具有较强的广谱抗细菌活性,对肺炎链球菌、流感嗜血菌的MIC为0.125μg/mL,尤其具有显著的抗MRSA的活性,对MRSA     

The depsidones from marine sponge-derived fungus Aspergillus unguis IB151 as an anti-MRSA agent: Molecular docking,pharmacokinetics analysis,and molecular dynamic simulation studies

Methicillin-resistant Staphylococcus aureus (MRSA) is an emerging nosocomial pathogen among hospitalized patients, with high morbidity and mortality rates. The discovery of a novel antibacterial is urgently needed to address this resistance problem. The present study aims to explore the antibacterial potential of three depsidone compounds: 2-clorounguinol (1), unguinol (2), and nidulin (3), isolated from the marine sponge-derived fungus Aspergillus unguis IB1, both in vitro and in silico. The antibacterial activity of all compounds was evaluated by calculating the Minimum inhibitory concentration (MIC) and Minimum bactericidal concentration (MBC) against MRSA using agar diffusion and total plate count methods, respectively. Bacterial cell morphology changes were  studied for the first time using scanning electron microscopy (SEM). Molecular docking, pharmacokinetics analysis, and molecular dynamics simulation were performed to determine possible protein–ligand interactions and the stability of the targeting penicillin-binding protein 2a (PBP2a) against 2-clorounguinol (1). The research findings indicated that compounds 1 to 3 exhibited MIC and MBC values of 2 µg/mL and 16 µg/mL against MRSA, respectively. MRSA cells displayed a distinct shape after the addition of the depsidone compound, as observed in SEM. According to the in silico study, 2-chlorounguinol exhibited the highest binding-free energy (BFE) with PBP2a (-6.7 kcal/mol). For comparison, (E)-3-(2-(4-cyanostyryl)-4-oxoquinazolin-3(4H)-yl) benzoic acid inhibits PBP2a with a BFE less than −6.6 kcal/mol. Based on the Lipinski's rule of 5, depsidone compounds constitute a class of compounds with good pharmacokinetic properties, being easily absorbed and permeable. These findings suggest that 2-chlorounguinol possesses potential antibacterial activity and could be developed as an antibiotic adjuvant to reduce antimicrobial resistance.     

中国南海紫柳珊瑚中倍半萜成分的研究

目的 研究南海紫柳珊瑚Muriceopsis flavida的化学成分。方法 应用正相硅胶、Sephadex LH-20凝胶、半制备RP-HPLC等多种色谱手段进行分离纯化,根据化合物的理化性质及波谱数据对化合物结构进行鉴定。采用琼脂扩散试验方法对化合物进行体外抗微生物活性测试。结果 从南海紫柳珊瑚中分离得到2个愈创木烷内酯型倍半萜,分别鉴定为menverins C(1)及D(2)。体外活性测试表明这两个化合物有不同程度的抗微生物活性,它们对巨大芽孢杆菌及微藻分别显示相当强的生长抑制作用。结论 化合物1, 2为首次从该属珊瑚中分离得到。Menverins C及D所表现出来的抗微生物作用,说明这些化合物在紫柳珊瑚的化学生态防御过程中可能具有重要的生态学功能。     

Pharmaceutical significance of Leuconostoc mesenteroides KS-TN11 isolated from Nile Tilapia,Oreochromis niloticus

Aquatic animals are known for their myriad of beneficial bacteria with diverse biologically active compounds. The current study was aimed to isolate and characterize potentially beneficial lactic acid bacteria from Nile Tilapia and evaluate their pharmaceutical applications. The fish samples were dissected and stomach, intestine, and gills were collected and serially diluted for the isolation of lactic acid bacteria (LAB) on BCP agar media. Identification of isolate was carried by biochemical and molecular characterization using API kit and 16S rRNA gene sequencing analysis, respectively. Further, KS-TN11 was assessed for α-glucosidase inhibitory potential using the chromogenic method. A lactic acid bacterium KS-TN11 was isolated from the stomach of Nile Tilapia and identified as Leuconostoc mesenteroides. Effect of KS-TN11 on lipid accumulation in adipocytes was done by using Oil Red O staining. The isolate showed strong antibacterial activity against a number of pathogenic bacteria in vitro. In addition, L. mesenteroides KS-TN11 KS-TN11 (50?mg/ml and 100?mg/ml) tends to inhibit adipogenesis in 3T3-L1 adipocytes and thus may have possible anti-obesity effects. Moreover, L. mesenteroides KS-TN11 exhibited substantial α–glucosidase inhibitory activities by 41.33% at 50?mg/ml and 64% at 100?mg/ml, respectively. The bacterium showed potent antibacterial activity against a number of pathogenic bacteria; in addition to alpha-glucosidase activity, and inhibition of lipid accumulation in 3T3-L1 cell line. These results reinforce KS-TN11 as a novel bacterium with an impending pharmaceutical application.     

Aspergillus flavus Growth Inhibition and Aflatoxin B1 Decontamination by Streptomyces Isolates and Their Metabolites

Aflatoxin B₁ is a potent carcinogen produced by Aspergillus flavus, mainly during grain storage. As pre-harvest methods are insufficient to avoid mycotoxin presence during storage, diverse curative techniques are being investigated for the inhibition of fungal growth and aflatoxin detoxification. Streptomyces spp. represent an alternative as they are a promising source of detoxifying enzymes. Fifty-nine Streptomyces isolates and a Streptomyces griseoviridis strain from the commercial product Mycostop^®, evaluated against Penicillium verrucosum and ochratoxin A during previous work, were screened for their ability to inhibit Aspergillus flavus growth and decrease the aflatoxin amount. The activities of bacterial cells and cell-free extracts (CFEs) from liquid cultures were also evaluated. Fifty-eight isolates were able to inhibit fungal growth during dual culture assays, with a maximal reduction going down to 13% of the control. Aflatoxin-specific production was decreased by all isolates to at least 54% of the control. CFEs were less effective in decreasing fungal growth (down to 40% and 55% for unheated and heated CFEs, respectively) and aflatoxin-specific production, with a few CFEs causing an overproduction of mycotoxins. Nearly all Streptomyces isolates were able to degrade AFB₁ when growing in solid and liquid media. A total degradation of AFB₁ was achieved by Mycostop^® on solid medium, as well as an almost complete degradation by IX20 in liquid medium (6% of the control). CFE maximal degradation went down to 37% of the control for isolate IX09. The search for degradation by-products indicated the presence of a few unknown molecules. The evaluation of residual toxicity of the tested isolates by the SOS chromotest indicated a detoxification of at least 68% of AFB₁’s genotoxicity.     

Chemical composition, antimicrobial and antioxidant activities of essential oil from Gnaphlium affine

The chemical composition of the essential oil from Gnaphlium affine was determined, and its antimicrobial and antioxidant activities were evaluated. Twenty-four compounds, representing 94.95% of the amount of total oil, were identified by gas chromatography-mass spectrometry (GC-MS) analysis. Main constituents of the essential oil were found to be eugenol (18.24%), linalool (10.62%), trans-caryophyllene (8.86%), α-terpineol (5.97%), p-cymene (5.75%), hexadecanoic acid (5.63%), γ-cadinene (4.98%), δ-cadinene (4.22%), α-humulene (3.22%), and (−)-β-elemene (3.15%). The essential oil revealed a remarkable antimicrobial effect against the tested food-borne microorganisms with the MIC and MBC values in the ranges of 0.2-1.56 μg/ml and 0.39-3.13 μg/ml, respectively. The essential oil showed a potent antioxidant activity in ABTS radical scavenging, lipid peroxidation and reducing power assay. It was suggested that the essential oil from G. affine may be a new potential source as natural antimicrobial and antioxidant agents applied in food systems.     

Antimicrobial activity of new 4,6-disubstituted pyrimidine, pyrazoline, and pyran derivatives

A number of new 2,6-didisubstituted pyrimidine, pyrazoline, and pyran derivatives were synthesized starting from their chalcone derivative. The synthesized compounds displayed different degrees of antimicrobial activity against Bscillus subtilis (Gram-positive), Pseudomonas aeruginosa (Gram-negative), and Streptomyces species (Actinomycetes).     

Characterization of the natural products in cocoyam (Colocasia esculenta) using GC–MS

Context: There is paucity of information in literature on the natural products in cocoyam [Colocasia esculenta Linn (Araceae)] that confer it with biological properties.

Objectives: This study investigated the antioxidant properties of C. esculenta and also reported for the first time the natural products in C. esculenta that justify its biological properties.

Materials and methods: The antioxidant activity of the methanol extract (50–250?μg/mL) of C. esculenta was determined using the 2,2-diphenyl-1-picryl hydrazyl (DPPH) radical and reducing power assays. Characterization of the natural products in C. esculenta was done using the gas chromatographic–mass spectrometric (GC–MS) technique. The experiment lasted for 3 months.

Results: GC–MS analysis of methanol/chloroform extract of the flour of C. esculenta indicated the presence of eight compounds, namely hexadecanoic acid methyl ester (0.43%), octadecanoic acid (20.91%), 9,12-octadecadienoyl chloride (0.77%), 11-octadecenoic acid methyl ester (2.12%), 9-octadecenoic acid (64.37%), 3-hexadecyloxycarbonyl-5-(2-hydroxylethyl)-4-methylimidazolium(1.36%), hexanedioic acid, bis(2-ethylhexyl)ester (1.36%) and 3,5-di-t-butyl phenol (3.27%). The total phenolic content of C. esculenta was 15.15?±?0.35?mg Gallic Acid Equivalence/g and it was significantly higher (p?<?0.05) than the total flavonoid (8.50?±?0.42?mg Quercetin Equivalence/g) and condensed tannin (4.40?±?0.14?mg Catechin Equivalence/g) contents, respectively. C. esculenta possessed strong antioxidant capacity though it was lower than that of standard quercetin.

Discussion and conclusion: Results showed that C. esculenta possesses strong antioxidant activity and also contains some important bioactive compounds that justify its medicinal properties as used in ethno-medicine.     

大青叶脂溶性成分的GC-MS分析

目的：研究大青叶药材的脂溶性成分.方法：用索氏提取法提取大青叶的脂溶性成分,甲酯化处理后,采用气相色谱-质谱联用技术首次对大青叶的脂溶性成分进行分析和鉴定.结果：从大青叶中鉴定了34个化合物,占大青叶脂溶性成分的73.98％,大青叶主要的脂溶性成分是α-亚麻酸（12.02％）,γ-谷甾醇（5.93％）,棕榈酸（5.6％）,7,10,13-二十碳三烯酸（4.82％）和油酸（3.78％）.结论：亚麻酸、棕榈酸和油酸是大青叶药材中的活性成分.     

4-Thiazolidinone and 1-thia-3,4,9-triaza fluorene conjugates: synthesis, characterization and antimicrobial screening

Some novel 4-thiazolidinone derivatives have been synthesized by the condensation of isatin/5-chloroisatin with thiosemicarbazide to yield thiosemicarbazones, which were then cyclized to form corresponding thia-3,4,9-triaza-fluoren-2-ylamines. These were reacted with substituted aldehydes to give corresponding Schiff bases, which were cyclized using thioglycolic acid in the presence of zinc chloride to obtain the 4-thiazolidinone derivatives. All the synthesized compounds were characterized by spectral (IR, MS and NMR) and elemental analysis. The compounds were screened for their antibacterial activity against Gram-positive bacteria (B. subtilis, S. aureus, B. pumilus and M. luteus), Gram-negative bacteria (P. aeruginosa, E. coli and P. fluorescens) and for antifungal activity against A. niger and P. chrysogenum by agar-diffusion method. The minimum inhibitory concentrations of these compounds were also determined by tube dilution method. The antimicrobial effectiveness of all the compounds was found to be concentration dependent. Two compounds—2-methyl-3-(1-thia-3, 4, 9-triaza-fluoren-2-yl)-thiazolidin-4-one (7aI) and 2-naphthalen-1-yl-3-(1-thia-3, 4, 9-tri aza-fluoren-2-yl)-thiazolidin-4-one (7aII)—exhibited good antibacterial activity. The antibacterial activity of all the compounds was found to be better than the antifungal activity.