Biodegradation of polyvinyl chloride, polystyrene, and polylactic acid microplastics in Tenebrio molitor larvae: Physiological responses.

It is widely understood that microplastics (MPs) can induce various biological stresses in macroinvertebrates that are incapable of biodegrading plastics. However, the biodegradation and physiological responses of plastic-degrading macroinvertebrates toward MPs of different degradability levels remain unexplored. In this study, Tenebrio molitor larvae (mealworms) were selected as a model of plastics-degrading macroinvertebrate, and were tested against three common plastics of different degradability rankings: polyvinyl chloride (PVC), polystyrene (PS), and polylactic acid (PLA) MPs (size <300 µm). These three MPs were biodegraded with the rate sequence of PLA > PS > PVC, resulting in a reversed order of negative physiological responses (body weight loss, decreased survival, and biomass depletion) of mealworms. Simultaneously, the levels of reactive oxygen species (ROS), antioxidant enzyme activities, and lipid peroxidation were uniformly increased as polymer degradability decreased and intermediate toxicity increased. PVC MPs exhibited higher toxicity than the other two polymers. The oxidative stresses were effectively alleviated by supplementing co-diet bran. The T. molitor larvae fed with PLA plus bran showed sustainable growth without an increase in oxidative stress. The results provide new insights into the biotoxicity of MPs on macroinvertebrates and offer comprehensive information on the physiological stress responses of plastic-degrading macroinvertebrates during the biodegradation of plastics with different degradability levels.

Biological Functions and Cross-Kingdom Host Gene Regulation of Small RNAs in Lactobacillus plantarum-Derived Extracellular Vesicles.

Extracellular vesicle-mediated transfer of microRNAs is a novel mode of cell-to-cell genetic transmission. Extracellular vesicles produced by microbes have been shown to contain significant quantities of physiologically active molecules such as proteins, lipids, and RNA, which could be transported to host cells and play a key role in both inter-kingdom signaling and physiological responses. In this study, we identified sRNAs by sequencing small RNAs (sRNAs) from Lactobacillus plantarum-derived extracellular vesicles (LDEVs) and detected the expression levels of vesicular sRNAs using quantitative reverse transcription-polymerase chain reaction (RT-PCR), which demonstrated the presence of microRNA-sized RNAs (msRNAs) within these vesicles. We chose sRNA71, a highly expressed msRNA, for further investigation, predicted its potential target genes for the human genome, and indicated that it could be translocated into mammalian cells. The biological functions of this sRNA71 were subsequently explored through cellular proteomics, western blot, and luciferase reporter assay. According to the findings, transfection with synthetic sRNA71 mimics substantially reduced Tp53 expression in HEK293T cells and suppressed the gene expression through binding to the 3' UTR of Tp53 mRNA. In conclusion, it is hypothesized that microbial-derived extracellular vesicles serve as carriers of functional molecules such as sRNAs, which play an essential role in regulating microbial-host communication.

Characterization of three different types of extracellular vesicles and their impact on bacterial growth.

Recently body fluids have been found to contain a class of nanoparticles released from cells, referred to as extracellular vesicles; exosomes are a type of small-diameter extracellular vesicle. We selected three types of sample: milk-derived exosomes, adipose-derived stem cell exosomes, and nanoparticles extracted from coconut water, to investigate their morphology, particle size distribution, protein content, and microRNA expression levels. Among the vesicles investigated, coconut nanoparticles had the greatest size distribution, and the protein content of coconut nanoparticles differed from that of mammalian exosomes. Using fluorescence microscopy, we determined that DiI-labeled extracellular vesicles could be absorbed by bacteria. Prominently, milk-derived exosomes could promote the growth of Escherichia coli K-12 MG1655 and Lactobacillus plantarum WCFS1. The studied extracellular vesicles could alter bacterial gene expression. Overall, this study identified differences in exogenous extracellular vesicles from different sources and revealed their supportive effects on microbial growth to make better utilization of microbial resources.

Isolation of Exosome-Like Nanoparticles and Analysis of MicroRNAs Derived from Coconut Water Based on Small RNA High-Throughput Sequencing.

In this study, the presence of microRNAs in coconut water was identified by real-time polymerase chain reaction (PCR) based on the results of high-throughput small RNA sequencing. In addition, the differences in microRNA content between immature and mature coconut water were compared. A total of 47 known microRNAs belonging to 25 families and 14 new microRNAs were identified in coconut endosperm. Through analysis using a target gene prediction software, potential microRNA target genes were identified in the human genome. Real-time PCR showed that the level of most microRNAs was higher in mature coconut water than in immature coconut water. Then, exosome-like nanoparticles were isolated from coconut water. After ultracentrifugation, some particle structures were seen in coconut water samples using 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate fluorescence staining. Subsequent scanning electron microscopy observation and dynamic light scattering analysis also revealed some exosome-like nanoparticles in coconut water, and the mean diameters of the particles detected by the two methods were 13.16 and 59.72 nm, respectively. In conclusion, there are extracellular microRNAs in coconut water, and their levels are higher in mature coconut water than in immature coconut water. Some exosome-like nanoparticles were isolated from coconut water, and the diameter of these particles was smaller than that of animal-derived exosomes.

Effects of microwave on extracellular vesicles and microRNA in milk.

This study demonstrates the effects of microwaves on the microRNA (miRNA) content of milk and milk extracellular vesicles (EV). We determined the miRNA concentration in milk subjected to different treatments using real-time PCR and a spectrophotometer. The miRNA expression and total RNA content of the microwaved milk samples were lower when compared with untreated milk. We measured the microstructure and the size distribution by scanning electron microscopy and dynamic light scattering to verify the loss of miRNA in microwaved milk due to damage to the EV. The results revealed that 2 different-sized EV were present and had an average size of 147.50 and 22.14 nm, respectively. Furthermore, acridine orange staining showed that the total RNA content in microwaved milk EV was lower than that in cow milk. These results suggest that EV may confer the protection and the stability of the miRNA in milk.

Fermentation Results in Quantitative Changes in Milk-Derived Exosomes and Different Effects on Cell Growth and Survival.

The discovery of microRNAs encapsulated in milk-derived exosomes has revealed stability under extreme conditions reflecting the protection of membranes. We attempted to determine the variations in nanoparticles derived from milk after fermentation, and provide evidence to determine the effects of these exosomes on cells with potential bioactivity. Using scanning electron microscopy and dynamic light scattering, we compared the morphology and particle size distribution of exosomes from yogurt fermented with three different combinations of strains with those from raw milk. The protein content of the exosome was significantly reduced in fermented milk. The cycle threshold showed that the expression of miR-29b and miR-21 was relatively high in raw milk, indicating a loss of microRNA after fermentation. Milk-derived exosomes could promote cell growth and activate the mitogen-activated protein kinase pathway. These findings demonstrated biological functions in milk exosomes and provided new insight into the nutrient composition of dairy products.

A new curvularin glycoside and its cytotoxic and antibacterial analogues from marine actinomycete Pseudonocardia sp. HS7.

Five curvularin macrolides (1-5) were isolated from the cultured broth of marine actinomycete Pseudonocardia sp. HS7 that was obtained from the cloacal aperture of sea cucumber Holothuria moebii. The structures of these isolates were characterized as (11S,15R)-11-hydroxycurvularin (1), (11R,15R)-11-hydroxycurvularin (2), curvularin-7-O-α-D-glucopyranoside (3), trans-dehydrocurvularin (4) and curvularin (5) based on their NMR and HRESIMS data as well as chemical degradation. Compound 3 is a new macrolide with a rare α-D-glucopyranose substituent. Compounds 1-4, 5a and 5c (the acyl products of 5), suppressed the proliferation of all six tested cancer cell lines and 4 is the most active compound with IC50 values ranging from 0.59 to 3.39 µM. The 11-hydroxycurvularins 1 and 2 also showed antibacterial activity inhibiting the growth of Escherichia coli.

Cytotoxic and anti-colorectal tumor effects of sulfated saponins from sea cucumber Holothuria moebii.

BACKGROUND: Whether sulfated saponins from Holothuria moebii inhibit the proliferation of colorectal cancer cells and have anti-colorectal tumor effects in animal model has not been investigated. PURPOSE: To evaluate the cytotoxic and anti-colorectal tumor effects of sulfated saponins from sea cucumber Holothuria moebii. METHOD: (1) Column chromatography was used to prepare the total and individual saponins and HPLC was applied to define the components of the total saponins; (2) the activity of the total and individual saponins inhibiting the proliferation of human colorectal cancer cells was determined by SRB assay and the apoptosis induced by the saponins was qualified using cytometric analysis with Annexin V-FITC/PI double staining; and (3) the antitumor effects of the sulfated saponins on colorectal CT-26 tumor-bearing Balb/c mice were tested. RESULTS: The total and individual sulfated saponins significantly inhibited the proliferation of four different human colorectal cancer cells with IC50 values ranging from 1.04 to 4.08 µM (or 1.46 to 3.24 µg/ml for total saponins) and induced late apoptosis at an early treatment time in cancer cells. The total saponins (120 mg/kg) had antitumor activity in colorectal CT-26 tumor-bearing Balb/c mice. CONCLUSION: The sulfated saponins from H. moebii remarkably inhibited the proliferation of different human colorectal cancer cells and had significant anti-colorectal tumor activity in animal model.

Synthesis and bioactivity of tripolinolate A from Tripolium vulgare and its analogs.

A new coniferol derivative, named as tripolinolate A (1), and 11 known compounds (2-12) were isolated from whole plants of Tripolium vulgare Nees. The structure of this new compound was determined as 4-(2S-methylbutyryl)-9-acetyl-coniferol based on its NMR and HRESIMS spectral analyses. A simple and efficient method was designed to prepare tripolinolate A and its 19 analogs including nine new chemical entities for bioactive assay. Tripolinolate A and its analog 4,9-diacetyl-coniferol were found to be the two most active compounds that significantly inhibited the proliferation of different cancer cell lines with IC50 values ranging from 0.36 to 12.9µM and induced apoptosis in tumor cells. Structure-activity relationship analysis suggested that the molecular size of acyl moieties at C-4 and C-9 position might have an effect on the activity of this type of coniferol derivatives.

Bioactive sulfated saponins from sea cucumber Holothuria moebii.

The bioactive ingredients of sea cucumber Holothuria moebii were investigated, and four sulfated saponins (1-4) and one desulfated saponin (3B) with an unusual 3,4-epoxy xylose were obtained from this study. Compound 2 is a new triterpenoid saponin and 3B is a new artificial compound. On the basis of the extensive NMR and HRESIMS data, their structures were assigned as 3-O-[ß-D-quinovopyranosyl-(1 → 2)-4-sodium sulfato-ß-D-xylopyranosyl]-25-acetoxy-22-oxo-9(11)-holostene-3ß,12α,17α-triol (2) and 3-O-[ß-D-quinovopyranosyl-(1 → 2)-3,4-epoxy-ß-xylopyranosyl]-22,25-epoxy-9(11)-holostene-3ß,12α,17α-triol (3B). Compounds 1-4 showed activity suppressing the proliferation of four different glioma cells with IC50 values ranging from 0.99 to 8.64 µM. New saponin 2 significantly induced apoptosis in human glioblastoma U87-MG cells and reduced the expression levels of several glioma metabolic enzymes of glycolysis and glutaminolysis. This study reveals for the first time that selectively targeting multiple glioma metabolic regulators of glycolysis and glutaminolysis might be one of the anti-glioma mechanisms of saponin 2.

Bioactive triterpenoid saponins and phenolic compounds against glioma cells.

A total of 54 natural origin compounds were evaluated for their activity in inhibiting the proliferation of glioma cells. Results showed that four Aesculus polyhydroxylated triterpenoid saponins (3-6), six Gleditsia triterpenoid saponins (7-12), and five phenolic compounds (43-46, 51) had dose-dependent activity suppressing the proliferation of both C6 and U251 cells. Structure-activity relationship analysis suggested that the acetyl group at C-28 for the Aesculus saponins and the monoterpenic acid moiety for the Gleditsia saponins could be critical for the activity of these active compounds. Aesculioside H (4), gleditsioside A (7), and feuric acid 3,4-dihydroxyphenethyl ester (FADPE, 46) were the three most active compounds from the different types of the active compounds and induced apoptosis and necrosis in glioma cells.

Polyoxygenated 24,28-epoxyergosterols inhibiting the proliferation of glioma cells from sea anemone Anthopleura midori.

Eleven sterols (1-11) and one carotenoid (12) were isolated and identified from sea anemone Anthopleura midori. Compounds 1-6 are rare polyoxygenated ergosterols with a 24,28-epoxy moiety. The structures of these epoxyergosterols were determined by NMR and HRESIMS analyses as well as their chemical-physical properties. Epoxyergosterols 1 and 2 were found to be new natural products and 3-6 are new compounds. Bioactive assay showed that compounds 1, 2, 3, 5, 7, 8, 11, and 12 inhibited the proliferation of rat glioma C6 and human glioma U251 cells with IC50 in a range of 2.41-80.45 µM. Further investigation suggested that 1 and 3 induced apoptosis in glioma cells and 1 blocked U251 cells at the G0/G1 phase.

Fatsioside A, a rare baccharane-type glycoside inhibiting the growth of glioma cells from the fruits of Fatsia japonica.

A novel baccharane-type triterpenoid glycoside named fatsioside A (1), together with ten oleanane glycosides, were isolated from the fruits of Fatsia japonica. The structure of fatsioside A was assigned as 3ß,15α,18α-trihydroxy-18,19-secolupane-12,19-dione 3-O-ß-D-glucopyranosyl-(1 → 2)-ß-D-glucopyranoside by extensive NMR and HRESIMS analyses. F. japonica is the third baccharane glycoside-containing species reported to date in the plant kingdom, while fatsioside A represents the first baccharane glycoside found in the Araliaceae family. Fatsioside A inhibited the growth of rat glioma C6 cells and human glioma U251 cells with IC50 values of 33.48 ± 2.01 µM and 77.58 ± 6.19 µM, respectively. Further investigation indicated that fatsioside A induced apoptosis and necrosis in glioma cells, and arrested the cell cycle at the G0/G1 phase.

Quantitative determination of triterpenoid glycosides in Fatsia japonica Decne. & Planch. using high performance liquid chromatography.

Fatsia japonica Decne. & Planch. is a triterpenoid glycoside-rich herb with anti-inflammatory activity for the treatment of rheumatoid arthritis. A method for quantitative analysis of the complex triterpenoid glycosides in this medicinal plant has not been established so far. In this study, a high performance liquid chromatography (HPLC) method was developed for simultaneous qualification of 11 glycosides in F. japonica. The analysis was performed on an ODS-2 Hypersil column (250mm×4.6mm, 5µm) with a binary gradient mobile phase of water and acetonitrile. The established HPLC method was validated in terms of linearity, sensitivity, stability, precision, accuracy, and recovery. Results showed that this method had good linearity with R(2) at 0.99992-0.99999 in the test range of 0.04-9.00µg/µL. The limit of detection (LOD) and limit of quantification (LOQ) for the standard compounds were 0.013-0.020µg/µL and 0.040-0.060µg/µL. The relative standard deviations (RSDs%) of run variations were 0.83-1.40% for intra-day and 0.84-3.59% for inter-day. The analyzed compounds in the samples were stable for at least 36h, and the spike recoveries of the detected glycosides were 99.67-103.11%. The developed HPLC method was successfully applied for the measurements of the contents of 11 triterpenoid glycoside in different parts of F. japonica. Taken together, the HPLC method newly developed in this study could be used for qualitative and quantitative analysis of the bioactive triterpenoid glycosides in F. japonica and its products.

Antitumor activity of caffeic acid 3,4-dihydroxyphenethyl ester and its pharmacokinetic and metabolic properties.

Caffeic acid 3,4-dihydroxyphenethyl ester (CADPE), a natural polyphenol from Sarcandra glabra, has potent in vitro anticancer activity through multiple targets. This study investigated its in vivo anticancer efficacy and its pharmacokinetic and metabolic characteristics. CADPE at any of the dosage regimes (ip 2.5 mg/kg at an interval of 7 h, 12 h, or 24 h for eight days) significantly decreased tumor growth in hepatoma H22 and sarcoma S180 tumor-bearing mice. CADPE also significantly inhibited H22-induced acute ascites development. The in vivo anticancer efficacies of CADPE in these tumor models were equivalent to those of 5-fluorouracil (10 mg/kg, ip) and cyclophosphamide (10 mg/kg, ip), and CADPE did not show any toxicity. A high performance liquid chromatography method with the aid of liquid chromatography/mass spectrometry was established and validated for the pharmacokinetic and metabolic studies of CADPE. CADPE was detected in blood and the organs including liver, kidney, heart, spleen, and brain 1 min after tail intravenous administration, indicating that CADPE was able to quickly distribute to these organs. CADPE was quickly hydrolyzed both in mice and in vitro mice plasma, but was much stable in vitro human plasma, suggesting a better bioavailability of CADPE in human than in mice. The major metabolites of CADPE in mice were caffeic acid, hydroxytyrosol, and a CADPE glucuronide. This was the first time to reveal the pharmacokinetic and metabolic characteristics of CADPE. Taken together, CADPE had potent in vivo antitumor activity and was able to rapidly reach the body organs and to be hydrolyzed in blood to anticancer agents of caffeic acid and hydroxytyrosol. This study suggested that CADPE has the potential for the treatment of cancers and is worthy of further study.

New capoamycin-type antibiotics and polyene acids from marine Streptomyces fradiae PTZ0025.

Capoamycin-type antibiotics (2-5) and polyene acids (6, 7) were isolated from marine Streptomyces fradiae strain PTZ0025. Their structures were established by extensive nuclear magnetic resonance (NMR) and high resolution electron spray ionization mass spectroscopy (HRESIMS) analyses and chemical degradation. Compounds 3, 4, 6, 7 were found to be new and named as fradimycins A (3) and B (4), and fradic acids A (6) and B (7). Compounds 3-5 showed in vitro antimicrobial activity against Staphylococcus aureus with a minimal inhibitory concentration (MIC) of 2.0 to 6.0 µg/mL. Interestingly, Compounds 3-5 also significantly inhibited cell growth of colon cancer and glioma with IC50 values ranging from 0.13 to 6.46 µM. Fradimycin B (4), the most active compound, was further determined to arrest cell cycle and induce apoptosis in tumor cells. The results indicated that fradimycin B (4) arrested the cell cycle at the G0/G1 phase and induced apoptosis and necrosis in colon cancer and glioma cells. Taken together, the results demonstrated that the marine natural products 3-5, particularly fradimycin B (4), possessed potent antimicrobial and antitumor activities.