Dae-Geun Song

Optimization of pressurized liquid extraction of carotenoids and chlorophylls from Chlorella vulgaris.

Pressurized liquid extraction (PLE) was applied to the extraction of carotenoids and chlorophylls from the green microalga Chlorella vulgaris. Four extraction techniques such as maceration (MAC), Soxhlet extraction (SOX), ultrasound assisted extraction (UAE), and PLE were compared, and both the extraction temperature (50, 105, and 160 degrees C) and the extraction time (8, 19, and 30 min), which are the two main factors for PLE, were optimized with a central composite design to obtain the highest extraction efficiency. The extraction solvent (90% ethanol/water) could adequately extract the functional components from C. vulgaris. PLE showed higher extraction efficiencies than MAC, SOX, and UAE. Temperature was the key parameter having the strongest influence on the extraction of carotenoids and chlorophylls from chlorella. In addition, high heat treatment (>110 degrees C) by PLE minimized the formation of pheophorbide a, a harmful chlorophyll derivative. These results indicate that PLE may be a useful extraction method for the simultaneous extraction of carotenoids and chlorophylls from C. vulgaris.

Inhibition of gastrointestinal lipolysis by green tea, coffee, and gomchui (Ligularia fischeri) tea polyphenols during simulated digestion.

Green tea, coffee, and gomchui (Ligularia fischeri) tea, which are rich in polyphenols, may exhibit antiobesity effects by inhibiting pancreatic lipase. However, the bioavailability of some polyphenols is poor due to either degradation or absorption difficulties in the gastrointestinal tract, thus making their beneficial effects doubtful. This study was conducted to evaluate the inhibitory effect of three beverages on lipolysis and the contribution of their major polyphenols during simulated digestion. During simulated digestion, gomchui tea was the most potent at inhibiting gastrointestinal lipolysis, whereas green tea was the least potent. The strongest lipase inhibitor among purified major polyphenols was a green tea polyphenol, (-)-epigallocatechin gallate (EGCG, IC(50) = 1.8 ± 0.57 μM), followed by di-O-caffeoylquinic acid isomers (DCQA, IC(50) from 12.7 ± 4.5 to 40.4 ± 2.3 μM), which are gomchui tea polyphenols. However, the stability of DCQA was greater than that of EGCG when subjected to simulated digestion. Taken together, gomchui tea, which has DCQA as the major polyphenol, showed stronger lipolysis inhibitory activity during simulated digestion compared to both green tea and coffee.

Rapid identification of furanocoumarins in Angelica dahurica using the Online LC-MMR-MS and their nitric oxide inhibitory activity in RAW 264.7 cells

INTRODUCTION Angelica dahurica (Fisch. Ex hoffm.) Benth. Et Hook. is a perennial herb that grows throughout Korea whose dried roots have been used to treat various diseases in Korean traditional medicine. The root extract contains diverse constituents, and it is necessary to determine the active compounds. OBJECTIVE To investigate the nitric oxide (NO) inhibitory activity in a root extract of A. dahurica and identify the most active compounds using LC-NMR-MS. METHODOLOGY In search of the anti-inflammatory constituents of A. dahurica extract, the HPLC-based activity profiling approach was used to investigate the extracts NO inhibitory activity. To directly identify the compounds, a hyphenated LC-NMR-MS technique was applied. Reversed-phase isocratic chromatography was performed using the acetonitrile-water solvent system on a C(30) column. The identification of the compounds was based on information from ESI/MS and 1H-NMR. RESULTS NO inhibitory activities for five main fractions of the extract were evaluated, which were identified by LC-NMR-MS as containing furanocoumarins: byakangelicol, oxypeucedanin, imperatorin, phellopterin and isoimperatorin. CONCLUSION The results obtained showed that the anti-inflammatory activities of A. dahurica could be linked to imperatorin and phellopterin.

A pepper MSRB2 gene confers drought tolerance in rice through the protection of chloroplast-targeted genes.

Background The perturbation of the steady state of reactive oxygen species (ROS) due to biotic and abiotic stresses in a plant could lead to protein denaturation through the modification of amino acid residues, including the oxidation of methionine residues. Methionine sulfoxide reductases (MSRs) catalyze the reduction of methionine sulfoxide back to the methionine residue. To assess the role of this enzyme, we generated transgenic rice using a pepper CaMSRB2 gene under the control of the rice Rab21 (responsive to ABA protein 21) promoter with/without a selection marker, the bar gene. Results A drought resistance test on transgenic plants showed that CaMSRB2 confers drought tolerance to rice, as evidenced by less oxidative stress symptoms and a strengthened PSII quantum yield under stress conditions, and increased survival rate and chlorophyll index after the re-watering. The results from immunoblotting using a methionine sulfoxide antibody and nano-LC-MS/MS spectrometry suggest that porphobilinogen deaminase (PBGD), which is involved in chlorophyll synthesis, is a putative target of CaMSRB2. The oxidized methionine content of PBGD expressed in E. coli increased in the presence of H2O2, and the Met-95 and Met-227 residues of PBGD were reduced by CaMSRB2 in the presence of dithiothreitol (DTT). An expression profiling analysis of the overexpression lines also suggested that photosystems are less severely affected by drought stress. Conclusions Our results indicate that CaMSRB2 might play an important functional role in chloroplasts for conferring drought stress tolerance in rice.

Effect of microfluidization on in vitro micellization and intestinal cell uptake of lutein from Chlorella vulgaris.

Chlorella is a nutrient-rich microalga that contains protein, lipid, minerals, vitamins, and high levels of lutein. This study evaluated the bioavailability of lutein from Chlorella vulgaris using a coupled in vitro digestion and human intestinal Caco-2 cell model. Lutein bioaccessibility was low, and approximately 75% of total C. vulgaris lutein was not micellized during the digestion process but remained in the insoluble digestate. Microfluidization improved lutein micellization efficiency during C. vulgaris digestion. C. vulgaris was microfluidized at a pressure exceeding 10000 psi, and the cell surface disruption was visualized by scanning electron microscopy. The mean C. vulgaris particle size was reduced from 3.56 to 0.35 μm with the microfluidization treatment. C. vulgaris microfluidization at 20000 psi was three times more efficient for aqueous lutein micelles production as compared with untreated C. vulgaris, and the final lutein content accumulated by intestinal Caco-2 cells was also higher with microfluidization. C. vulgaris lutein stability was not affected by microfluidization. These results indicate that microfluidization may be useful for improving lutein bioaccessibility from C. vulgaris during food processing.

Daurinol, a catalytic inhibitor of topoisomerase IIα, suppresses SNU-840 ovarian cancer cell proliferation through cell cycle arrest in S phase

Daurinol, a lignan from the ethnopharmacological plant Haplophyllum dauricum, was recently reported to be a novel topoisomerase II inhibitor and an alternative to the clinical anticancer agent etoposide based on a colorectal cancer model. In the present study, we elucidated the detailed biochemical mechanism underlying the inhibition of human topoisomerase IIα by daurinol based on a molecular docking study and in vitro biochemical experiments. The computational simulation predicted that daurinol binds to the ATP-binding pocket of topoisomerase IIα. In a biochemical assay, daurinol (10-100 µM) inhibited the catalytic activity of topo-isomerase IIα in an ATP concentration-dependent manner and suppressed the ATP hydrolysis activity of the enzyme. However, daurinol did not inhibit topoisomerase I activity, most likely because topoisomerase I does not contain an ATP-binding domain. We also evaluated the anti-proliferative activity of daurinol in ovarian, small cell lung and testicular cancer cells, common target cancers treated with etoposide. Daurinol potently inhibited SNU-840 human ovarian cancer cell proliferation through cell cycle arrest in S phase, while etoposide induced G2/M phase arrest. Daurinol induced the increased expression of cyclin E, cyclin A and E2F-1, which are important proteins regulating S phase initiation and progression. Daurinol did not induce abnormal cell and nuclear enlargement in SNU-840 cells, in contrast to etoposide. Based on these data, we suggest that daurinol is a potential anticancer drug candidate for the treatment of human ovarian cancer with few side effects.

Phlorofucofuroeckol-A, a potent inhibitor of aldo-keto reductase family 1 member B10, from the edible brown alga Eisenia bicyclis

Aldo-keto reductase family 1 member B10 (AKR1B10) belongs to a superfamily of NADPH-dependent aldo-keto reductases and is considered a biomarker of several cancers. Inhibition of recombinant human AKR1B10 (rhAKR1B10) was assayed using 31 seaweed extracts, among which, an Eisenia bicyclis extract was selected for further study. To identify the compounds in E. bicyclis responsible for inhibitory effects on rhAKR1B10, five compounds were isolated by bioactivity-guided fractionation and isolation. Among them, phlorofucofuroeckol-A (PFF-A), isolated from an ethyl acetate fraction, exhibited the greatest inhibition of rhAKR1B10. The inhibitory rate of PFF-A against rhAKR1B10 was 61.41% at 10 μM, with an IC50 of 6.22 μM. Enzyme kinetic analyses revealed non-competitive inhibition with a KD of 2.76 μM. These results indicate that PFF-A from E. bicyclis may be a promising anticancer agent.

Secretome Profiling Reveals the Signaling Molecules of Apoptotic HCT116 Cells Induced by the Dietary Polyacetylene Gymnasterkoreayne B

Dietary polyacetylenes from various foods have been receiving attention as promising cancer chemopreventive agents. However, until now, the detailed molecular mechanism and the regulatory proteins underlying these effects have not been elucidated. We investigated the effects of gymnasterkoreayne B (GKB), a model dietary polyacetylene from wild vegetables, on the programmed cell death of HCT116 human colorectal cancer cells. GKB inhibited HCT116 cell proliferation by inducing apoptotic cell death. GKB treatment resulted in ROS accumulation, leading to the activation of both intrinsic and extrinsic apoptotic pathway. We also found that FN1, TGFB1, APP, SERPINE1, HSPD1, SOD1, TXN, and ACTN4 may act as secretory signaling molecules during GKB-induced apoptotic cell death using LC-MS/MS identification followed by spectrum counting, statistical calculation, and gene ontology analysis. The secretory proteins suggested in this study may be promising candidates involved in apoptotic cell death of cancer cells induced by GKB that warrant further functional study.

Ranitidine-induced anaphylaxis: clinical features, cross-reactivity, and skin testing

Histamine H2 receptor antagonists are commonly prescribed medications and are known to be well tolerated. However, 99 cases of ranitidine‐induced anaphylaxis occurred in Korea from 2007 to 2014.

The three proline residues (P25, P242, and P434) of Agrobacterium CP4 5-enolpyruvylshikimate-3-phosphate synthase are crucial for the enzyme activity.

Multiple sequence alignments showed that the prolines at the 25th, 129th, 153rd, 242nd, 322nd, and 434th amino acids in 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) from Agrobacterium sp. strain CP4 are strongly conserved in various prokaryotic EPSPS proteins. Single point mutations of the conserved prolines to alanine (P25A, P153A, P242A, P322A, and P434A) were introduced in the CP4 EPSPS in order to investigate the importance of the conserved prolines for the enzyme properties. The point mutations caused decreases in substrate binding affinity and catalytic efficiency as well as the glyphosate resistance, in general. Especially, the 25th and 242nd prolines located in the polypeptide hinges connecting top and bottom domains of CP4 EPSPS as well as the 434th proline at the C-terminus of the enzyme turned out to be crucial for the enzyme activity.