Platelet extracellular vesicles are efficient delivery vehicles of doxorubicin, an anti-cancer drug: preparation and in vitro characterization.

Platelet extracellular vesicles (PEVs) are an emerging delivery vehi for anticancer drugs due to their ability to target and remain in the tumor microenvironment. However, there is still a lack of understanding regarding yields, safety, drug loading efficiencies, and efficacy of PEVs. In this study, various methods were compared to generate PEVs from clinical-grade platelets, and their properties were examined as vehicles for doxorubicin (DOX). Sonication and extrusion produced the most PEVs, with means of 496 and 493 PEVs per platelet (PLT), respectively, compared to 145 and 33 by freeze/thaw and incubation, respectively. The PEVs were loaded with DOX through incubation and purified by chromatography. The size and concentration of the PEVs and PEV-DOX were analyzed using dynamic light scattering and nanoparticle tracking analysis. The results showed that the population sizes and concentrations of PEVs and PEV-DOX were in the ranges of 120-150 nm and 1.2-6.2 × 1011 particles/mL for all preparations. The loading of DOX determined using fluorospectrometry was found to be 2.1 × 106, 1.7 × 106, and 0.9 × 106 molecules/EV using freeze/thaw, extrusion, and sonication, respectively. The internalization of PEVs was determined to occur through clathrin-mediated endocytosis. PEV-DOX were more efficiently taken up by MDA-MB-231 breast cancer cells compared to MCF7/ADR breast cancer cells and NIH/3T3 cells. DOX-PEVs showed higher anticancer activity against MDA-MB-231 cells than against MCF7/ADR or NIH/3T3 cells and better than acommercial liposomal DOX formulation. In conclusion, this study demonstrates that PEVs generated by PLTs using extrusion, freeze/thaw, or sonication can efficiently load DOX and kill breast cancer cells, providing a promising strategy for further evaluation in preclinical animal models. The study findings suggest that sonication and extrusion are the most efficient methods to generate PEVs and that PEVs loaded with DOX exhibit significant anticancer activity against MDA-MB-231 breast cancer cells.

What is the context?● Current synthetic drug delivery systems can have limitations and side effects.● Platelet extracellular vesicles (PEVs) are a natural and potentially safer alternative for delivering cancer drugs to tumors.● However, there is still a lack of understanding about how to produce PEVs and how effective they are in delivering drugs.What is new?● We compared different methods for producing PEVs from clinical-grade platelets and found that sonication and extrusion were the most effective methods.● The PEVs were loaded with a cancer drug called doxorubicin (DOX) and tested their ability to kill breast cancer cells.What is the impact?● PEVs loaded with DOX were effective at killing cancer cells, especially MDA-MB-231 breast cancer cells.● This study demonstrates that PEVs are a promising strategy for delivering cancer drugs to tumors and that sonication and extrusion are the most efficient methods for producing PEVs.● The results suggest that further evaluation of PEVs in preclinical animal models is warranted to determine their potential as a cancer drug delivery system.Abbreviations: ADP: adenosine diphosphate; bFGF: basic fibroblast growth factor; BSA: bovine serum albumin; CD41: platelet glycoprotein IIb; CD62P: P-selectin; CFDASE: 5-(and-6)-carboxyfluorescein diacetate: succinimidyl ester; CPLT: cryopreserved platelet; CPZ: chlorpromazine hydrochloride; CTC: circulating tumor cell; DMSO: dimethyl sulfoxide; DDS: drug delivery system; DOX: doxorubicin; EPR: enhanced permeability and retention; EV: extracellular vesicle; FBS: fetal bovine serum; GMP: good manufacturing practice; GF: growth factor; HER2: human epidermal growth factor receptor 2; HGF: hepatocyte growth factor; Lipo-DOX: liposomal doxorubicin; MDR: multi-drug resistance; MMP-2: matrix metalloproteinase-2; MP: microparticle; MSC: mesenchymal stromal cell; NP: nanoparticle; NTA: nanoparticle tracking analysis; PAR-1: protease activated receptor-1; PAS: platelet additive solution; PBS: phosphate-buffered saline; PC: platelet concentrate; PEG: polyethylene glycol; PEV: platelet-derived extracellular vesicle; DOX-PEV: doxorubicin-loaded platelet-derived extracellular vesicle-encapsulated; PFA: paraformaldehyde; PF4: platelet factor 4; P-gp: P-glycoprotein; PLT: platelet; PS: phosphatidylserine; SDS-PAGE: sodium dodecylsulfate polyacrylamide gel electrophoresis; SEM: scanning electron microscopy; TCIPA: tumor cell-induced PLT aggregation; TDDS: targeted drug delivery system; TEG: thromboelastography; TF: tissue factor; TF-EV: extracellular vesicle expressing tissue factor; TME: tumor microenvironment; TNBC: triple-negative breast cancer; TXA2: thromboxane-A2; VEGF: vascular endothelial growth factor; WHO: World Health Organization.

Design, structure-activity relationships, and enzyme kinetic studies of tricyclic and tetracyclic coumarin-based sulfamates as steroid sulfatase inhibitors.

Inhibition of steroid sulfatase (STS) decreases estrogen production and thus, suppresses tumor proliferation. Inspired by irosustat, the first STS inhibitor in clinical trials, we explored twenty-one tricyclic and tetra-heterocyclic coumarin-based derivatives. Their STS enzyme kinetic parameters, docking models, and cytotoxicity toward breast cancer and normal cells were evaluated. Tricyclic derivative 9e and tetracyclic derivative 10c were the most promising irreversible inhibitors developed in this study, with KI of 0.05 and 0.4 nM, and kinact/KI ratios of 28.6 and 19.1 nM-1min-1 on human placenta STS, respectively.

The Design, Structure-Activity, and kinetic studies of 3-Benzyl-5-oxa-1,2,3,4-Tetrahydro-2H-chromeno-(3,4-c)pyridin-8-yl sulfamates as Steroid sulfatase inhibitors.

Steroid sulfatase inhibitors block the local production of estrogenic steroids and are attractive agents for the treatment of estrogen-dependent cancers. Inspiration of coumarin-based inhibitors, we synthesized thirty-two 5-oxa-1,2,3,4-tetrahydro-2H-chromeno-(3,4-c)pyridin-8-yl sulfamates, focusing on the substitution derivatives on the adjacent phenyl ring and evaluated their abilities to block STS from human placenta and MCF-7 cells. SAR analysis revealed that the incorporation of chlorine at either meta and/or para position of the adjacent phenyl ring of the tricyclic skeleton enhanced STS inhibition. Di-substitutions at the adjacent phenyl ring were superior to mono and tri-substitutions. Further kinetic analysis of these compounds revealed that chloride-bearing compounds, such as 19m, 19v, and 19w, had KI of 0.02 to 0.11 nM and kinact/KI ratios of 8.8-17.5 nM-1min-1, a parameter indicated for the efficiency of irreversible inhibition. We also used the docking model to illustrate the difference in STS inhibitory potency of compounds. Finally, the safety and anti-cancer activity of selected compounds 19m, 19v, and 19w were also studied, showing the results of low cytotoxicity on NHDF cell line and being more potent than irosustat on ZR-75-1 cell, which was a hormone-dependent cancer cell line with high STS expression.

Anti-adipogenic activities of pheophorbide a and pyropheophorbide a isolated from wild bitter gourd (Momordica charantia L. var. abbreviata Seringe) in vitro.

BACKGROUND: The wild bitter gourd (WBG) is a commonly consumed vegetable in Asia that has antioxidant and hypoglycemic properties. The present study aimed to investigate the anti-adipogenic activities of isolated compounds from WBG on 8-day differentiated cultures of 3 T3-L1 adipocytes that were then stained with Oil Red O (ORO) or diamidino-2-phenylindole (DAPI). RESULTS: ORO stains of the methanol extracts of de-seeded HM86 cultivar of WBG (WBG-M) and the ethyl acetate fractions (WBG-M-EA) showed anti-adipogenic activities against differentiated adipocytes. Two chlorophyll-degraded compounds, pheophorbide a (1) and pyropheophorbide a (2), were isolated from WBG-M-EA. Treatments with 1 (5, 10, and 20 µmol L-1 ) and 2 (2.5, 5, and 10 µmol L-1 ) showed dose-dependent reductions in lipid accumulations and reduced nuclear DAPI stains in differentiated 3 T3-L1 adipocytes. The concentrations for 50% inhibition against lipid accumulations of 1 and 2, respectively, were 16.05 and 7.04 µmol L-1 . Treatments with 1 and 2 showed enhanced lactate dehydrogenase release in the first 4-day cell mitotic clonal expansions during the differentiating cultural processes, although the effect was less on the non-differentiating cultural processes. Thus, 1 and 2 were more toxic to differentiating adipocytes than to non-differentiated pre-adipocytes, which partly resulted in anti-adipogenic activities with lowered lipid accumulations. CONCLUSION: Both 1 and 2 showed anti-adipogenic activities in cell models. These chlorophyll-degraded compounds commonly exist in several vegetables during storage or edible seaweeds, which will provide resources for further investigations aiming to test anti-obesity in animal studies. © 2022 Society of Chemical Industry.

Reductions of copper ion-mediated low-density lipoprotein (LDL) oxidations of trypsin inhibitors, the sweet potato root major proteins, and LDL binding capacities.

BACKGROUND: The root major proteins of sweet potato trypsin inhibitors (SPTIs) or named sporamin, estimated for 60 to 80% water-soluble proteins, exhibited many biological activities. The human low-density lipoprotein (LDL) showed to form in vivo complex with endogenous oxidized alpha-1-antitrypsin. Little is known concerning the interactions between SPTIs and LDL in vitro. RESULTS: The thiobarbituric-acid-reactive-substance (TBARS) assays were used to monitor 0.1 mM Cu2+-mediated low-density lipoprotein (LDL) oxidations during 24-h reactions with or without SPTIs additions. The protein stains in native PAGE gels were used to identify the bindings between native or reduced forms of SPTIs or soybean TIs and LDL, or oxidized LDL (oxLDL). It was found that the SPTIs additions showed to reduce LDL oxidations in the first 6-h and then gradually decreased the capacities of anti-LDL oxidations. The protein stains in native PAGE gels showed more intense LDL bands in the presence of SPTIs, and 0.5-h and 1-h reached the highest one. The SPTIs also bound to the oxLDL, and low pH condition (pH 2.0) might break the interactions revealed by HPLC. The LDL or oxLDL adsorbed onto self-prepared SPTIs-affinity column and some components were eluted by 0.2 M KCl (pH 2.0). The native or reduced SPTIs or soybean TIs showed different binding capacities toward LDL and oxLDL in vitro. CONCLUSION: The SPTIs might be useful in developing functional foods as antioxidant and nutrient supplements, and the physiological roles of SPTIs-LDL and SPTIs-oxLDL complex in vivo will investigate further using animal models.

Design and synthesis of 3-benzylaminocoumarin-7-O-sulfamate derivatives as steroid sulfatase inhibitors.

Steroid sulfatase (STS) is a sulfatase enzyme that catalyzes the conversion of sulfated steroid precursors to free steroid. The inhibition of STS could abate estrogenic steroids that stimulate the proliferation and development of breast cancer, and therefore STS is a potential target for adjuvant endocrine therapy. In this study, a series of 3-benzylaminocoumarin-7-O-sulfamate derivatives targeting STS were designed and synthesized. Structure-relationship activities (SAR) analysis revealed that attachment of a benzylamino group at the 3-position of coumarin improved inhibitory activity. Compound 3j was found to have the highest inhibition activity against human placenta isolated STS (IC50  0.13 µM) and MCF-7 cell lines (IC50 1.35 µM). Kinetic studies found compound 3j to be an irreversible inhibitor of STS, with KI and kinact value of 86.9 nM and 158.7 min-1, respectively.

Design of Diarylheptanoid Derivatives as Dual Inhibitors Against Class IIa Histone Deacetylase and ß-amyloid Aggregation.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder with multiple etiologies. Beta-amyloid (Aß) self-aggregation and overexpression of class IIa histone deacetylases (HDACs) are strongly implicated with AD pathogenesis. In this study, a series of novel diarylheptanoid derivatives were designed, synthesized and evaluated for use as dual Aß self-aggregation and class IIa HDAC inhibitors. Among these compounds, 4j, 5c, and 5e displayed effective inhibitions for Aß self-aggregation, HDAC5 activity and HDAC7 activity with IC50 values of <10 µM. The compounds contain three common features: (1) a catechol or pyrogallol moiety, (2) a carbonyl linker and (3) an aromatic ring that can function as an HDAC cap and create hydrophobic interactions with Aß1-42. Furthermore, compounds 4j, 5c, and 5e showed no significant cytotoxicity to human neuroblastoma SH-SY5Y cells and also exhibited neuroprotective effect against H2O2-induced toxicity. Overall, these promising in vitro data highlighted compounds 4j, 5c, and 5e as lead compounds that are worthy for further investigation.

Antiglycation, radical scavenging, and semicarbazide-sensitive amine oxidase inhibitory activities of acetohydroxamic acid in vitro.

Advanced glycation endproducts (AGEs) can promote intracellular reactive oxygen species production, and the levels of AGEs are highly correlated with cardiovascular disease and diabetes complications. Acetohydroxamic acid (acetH) is a bacterial urease inhibitor drug used to treat kidney stones and infections in the urinary tract, and hydroxyurea (HU) is a drug used for antineoplasm and sickle cell diseases. Both acetH and HU are hydroxamic acid derivatives. It was found that acetH and HU at 2.5 or 5 mM showed anti-AGE formation by lowering the AGEs' fluorescent intensities and Nε-(carboxymethyl)lysine formation in bovine serum albumin/galactose models, and both showed better and significant differences (P<0.05) compared to the positive control of aminoguanidine. Regarding radical scavenging activities, the half-inhibition concentrations (IC50) of acetH against α,α-diphenyl-ß-picrylhydrazyl radical and hydroxyl radical were 34.86 and 104.42 µM, respectively. The IC50 of acetH against semicarbazide-sensitive amine oxidase was 10.56 µM, and acetH showed noncompetitive inhibition respective to the substrates (benzylamine). The antiglycation, antioxidant, and semicarbazide-sensitive amine oxidase inhibitory activities of acetH prove that it has the potential for treating cardiovascular disease and diabetes complications and it needs further investigation in animal models.

Hot-Water Extracts from Roots of Vitis thunbergii var. taiwaniana and Identified ε-Viniferin Improve Obesity in High-Fat Diet-Induced Mice.

In this study, hot-water extracts (HW) from roots of Vitis thunbergii var. taiwaniana (VTT-R) were shown to lower levels of lipid accumulation significantly (P < 0.01 or 0.001) compared to the control in 3T3-L1 adipocytes. The VTT-R-HW (40 mg/kg) interventions concurrent with a high-fat (HF) diet in C57BL/6 mice over a 5 eek period were shown to reduce body weights significantly (P < 0.05) compared to those of mice fed a HF diet under the same food-intake regimen. The (+)-ε-viniferin isolated from VTT-R-HW was shown to reduce the size of lipid deposits significantly compared to the control (P < 0.05 or 0.001) in 3T3-L1 adipocytes, and dose-dependent 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase inhibitions showed that the 50% inhibitory concentration was calculated to be 96 µM. The two-stage (+)-ε-viniferin interventions (10 mg/kg, day 1 to day 38; 25 mg/kg, day 39 to day 58) were shown to lower mice body weights significantly (P < 0.05 or 0.001), the weight ratio of mesenteric fat, blood glucose, total cholesterol, and low-density lipoprotein compared to that of the HF group under the same food-intake regimen but without concurrent VTT-R-HW interventions. It might be possible to use VTT-R-HW or (+)-ε-viniferin as an ingredient in the development of functional foods for weight management, and this will need to be investigated further.

Effects of hot-water extracts from Ganoderma lucidum residues and solid-state fermentation residues on prebiotic and immune-stimulatory activities in vitro and the powdered residues used as broiler feed additives in vivo.

BACKGROUND: Large amounts of Ganoderma lucidum (GL) commercial products are provided in the worldwide market such as powders, tea bags, or capsules as dietary supplements which contained triterpenoids and/or polysaccharides. Therefore, it was estimated that several thousand tons of GL residues (GLR) are produced and discarded. For recycling uses, the aim of this study was to evaluate the benefits of two hot-water extracts from GLR (HWP_GLR) and solid-state fermentation GLR inoculated with GL mycelia (HWP_GLRF) on the growths of Lactobacillus rhamnosus and Bifidobacterium longum. The RAW264.7 cells were used to investigate the effects of HWP_GLR and HWP_GLRF on nitric oxide productions, phagocytic activities against FITC-labeled E. coli, and to lower lipopolysaccharide (LPS)-binding capacities. The powders of GLR and GLRF were used as additives in the commercial feeds for feeding broiler chicks in vivo to evaluate the immune-stimulatory and prebiotic activities. RESULTS: HWP_GLR and HWP_GLRF with molecular size 5 to 8 kDa were showed to stimulate growths of L. rhamnosus and B. longum. It was found that in the presence of polymyxin B HWP_GLR and HWP_GLRF could stimulate nitric oxide productions, elevate phagocytic activities against FITC-labeled E. coli, and to lower lipopolysaccharide-binding capacities in RAW264.7 cells. The broiler chicks were selected for feedings in vivo. The 1-day-old chicks were fed commercial feeds for 1 week, and then were fed without or with 4 or 8 % of GLR and GLRF additives for 3 weeks. There was no significant weight difference among feeding groups. However, the phagocytosis and natural killer cytotoxicity in the peripheral bloods, and prebiotic activities of bifidobacteria in feces of GLR and/or GLRF groups were significantly different compared to the control (P < 0.05). CONCLUSIONS: The GLR, GLRF, and their hot-water extracts with beneficial activities could be processed as feed additives which could increase the waste-recycling.

Vasorelaxing and antihypertensive activities of synthesized peptides derived from computer-aided simulation of pepsin hydrolysis of yam dioscorin.

BACKGROUND: We reported that yam dioscorin and its peptic hydrolysates exhibited ACE inhibition and antihypertensive effects on SHRs, however, the active peptides are not really isolated until now. Using ACE inhibitory screenings, two penta-peptides, KTCGY and KRIHF, were selected for ex vivo and in vivo experiments. RESULTS: KTCGY, KRIHF, and captopril were shown to have similar vasodilating effects against phenylephrine (PE)-induced tensions in rat endothelium-dependent thoracic aortic rings, however, KTCGYKTCGY (two-repeated KTCGY) and TCGYTCGY (two-repeated TCGY) were showed endothelium-independent vasodilating effects against PE-induced tensions. KTCGY, KRIHF (10 or 20 mg/kg), and captopril (10 mg/kg) were used to evaluate antihypertensive activity during 24-h after a single oral administration to spontaneously hypertensive rats (SHRs). The KTCGY and KRIHF showed significantly different and reduced the systolic blood pressure of SHRs compared to the blank. CONCLUSIONS: These results suggest that KTCGY and KRIHF may contribute important roles in yam dioscorin for regulating blood pressure in vivo.

Inhibitory activities of acteoside, isoacteoside, and its structural constituents against protein glycation in vitro.

BACKGROUND: Advanced glycation end products (AGE) are substances that can induce insulin resistance in adipocyte, hepatocyte and muscle cells. This resistance correlates highly with cardiovascular disease and diabetic complications. Acteoside (A), a phenylethanoid glycoside, is an active compound in several plants and traditional herbal medicines. Acteoside, its structural isomer, isoacteoside (I), and their constituents, caffeic acid (C) and 3,4-dihydroxyphenylethanol (D), were used in the study to investigate the inhibitory activity against AGE formations in vitro. RESULTS: AGE formations were detected by anti-(Nϵ-(carboxymethyl)lysine (anti-CML), using bovine serum albumin (BSA)/glucose (glc) and BSA/galactose (gal) as models, or by anti-argpyrimidine (anti-AP), using BSA/methylglyoxal (MGO) as models. It was found that A, I, C, or D, each at 5 mM, could attenuate the CML formations detected by ELISA in the BSA/gal model of a 3-day or 5-day reaction, and showed significant differences (P < 0.01 or P < 0.001) compared to the control. However, these compounds showed a minor effect after a 7-day incubation. It was also found that C or D could lower the CML formations in the BSA/glc model and showed significant differences (P < 0.05 or P < 0.01) compared to the control after a 3-day, 5-day and 7-day reaction. It was found that A, I, C, or D, each at 0.5 mM or 5 mM, could attenuate the AP formations in the BSA/MGO model of a 3-day reaction and showed significant differences (P < 0.001) compared to the control. CONCLUSIONS: The results suggest the potential anti-glycation activities of A and I in vitro may apply to cell models at higher glucose concentrations or to diabetic animal models, and need further investigation.

Ethanolic extracts and isolated compounds from small-leaf grape (Vitis thunbergii var. taiwaniana) with antihypertensive activities.

This study aimed to investigate the antihypertensive effects of ethanolic extracts (EE) and compounds isolated from the small-leaf grape (Vitis thunbergii var. taiwaniana, VTT). The highest antiangiotensin-converting enzyme (anti-ACE) was found in stem-EE (IC50 was 69.5 µg/mL). In spontaneously hypertensive rats (SHRs), stem-EE effectively reduced blood pressure 24 h after administration of a single oral dose or when administered daily for 4 weeks. The isolated compounds, including (+)-vitisin A, ampelopsin C, and (+)-ε-viniferin, were shown to have anti-ACE and vasodilating effects against phenylephrine-induced tensions in an endothelium-intact aortic ring, with (+)-vitisin A being the most effective compound. Compared to control rats, SHRs showed significantly reduced systolic and diastolic blood pressures 24 h after a single oral dose of (+)-vitisin A (10 mg/kg) or captopril (2 mg/kg). These results suggest that the development of functional foods with VTT extracts may be beneficial for regulating blood pressure.

Biological activities and applications of dioscorins, the major tuber storage proteins of yam.

Yam tubers, a common tuber crop and an important traditional Chinese medicine in Taiwan, have many bioactive substances, including phenolic compounds, mucilage polysaccharides, steroidal saponins and proteins. Among the total soluble proteins, 80% of them are dioscorins. In the past two decades, many studies showed that dioscorins exhibited biological activities both in vitro and in vivo, including the enzymatic, antioxidant, antihypertensive, immunomodulatory, lectin activities and the protecting role on airway epithelial cells against allergens in vitro. Some of these activities are survived after chemical, heating process or enzymatic digestion. Despite of lacking the intact structural information and the detail action mechanisms in the cells, yam dioscorins are potential resources for developing as functional foods and interesting targets for food protein researchers.

Antioxidant and amine oxidase inhibitory activities of hydroxyurea.

Hydroxyurea (HU, NH(2)CONHOH), or hydroxycarbamide, is a hydroxamic acid derivative used as a drug for anti-neoplasm and sickle-cell disease. In this study, HU was found to have antioxidant activities against 2,2-diphenyl-1-picrylhydrazyl (DPPH) and hydroxyl radicals and dose-dependent inhibitory activities against monoamine oxidase (MAO)-A, MAO-B, and semicarbazide-sensitive amine oxidase (SSAO) as compared to controls of clorgyline, deprenyl, and semicarbazide respectively. HU showed mixed-type, competitive-type, and competitive-type inhibition, respectively, with respect to substrates of MAO-A, MAO-B, and SSAO with apparent inhibition constants (Ki) of 19.46, 5.38, and 1.84 microM.

Synthesis of N-hydroxycinnamides capped with a naturally occurring moiety as inhibitors of histone deacetylase.

Histone deacetylase (HDAC) inhibitors are regarded as promising therapeutics for the treatment of cancer. All reported HDAC inhibitors contain three pharmacophoric features: a zinc-chelating group, a hydrophobic linker, and a hydrophobic cap for surface recognition. In this study we investigated the effectiveness of osthole, a hydrophobic Chinese herbal compound, as the surface recognition cap in hydroxamate-based compounds as inhibitors of HDAC. Nine novel osthole-based N-hydroxycinnamides were synthesized and screened for enzyme inhibition activity. Compounds 9 d, 9 e, 9 g exhibited inhibitory activities (IC(50)=24.5, 20.0, 19.6 nM) against nuclear HDACs in HeLa cells comparable to that of suberoylanilide hydroxamic acid (SAHA; IC(50)=24.5 nM), a potent inhibitor clinically used for the treatment of cutaneous T-cell lymphoma (CTCL). While compounds 9 d and 9 e showed SAHA-like activity towards HDAC1 and HDAC6, compound 9 g was more selective for HDAC1. Compound 9 d exhibited the best cellular effect, which was comparable to that of SAHA, of enhancing acetylation of either alpha-tubulin or histone H3. Molecular docking analysis showed that the osthole moiety of compound 9 d may interact with the same hydrophobic surface pocket exploited by SAHA and it may be modified to provide class-specific selectivity. These results suggest that osthole is an effective hydrophobic cap when incorporated into N-hydroxycinnamide-derived HDAC inhibitors.

Antioxidant, anti-semicarbazide-sensitive amine oxidase, and anti-hypertensive activities of geraniin isolated from Phyllanthus urinaria.

The wrinkle-fruited leaf flower (Phyllanthus urinaria L.) (Euphorbiaceae) is widely used as a traditional folk medicine for inflammatory relief. Geraniin, the hydrolysable tannin, was purified by a series of chromatographic processes from the 70% aqueous acetone extracts of P. urinaria and identified by NMR [1H (500 MHz) and 13C NMR (126 MHz)] spectra and mass spectroscopy. The scavenging activities of geraniin against DPPH radicals (half-inhibition concentration, IC50, were 0.92 and 1.27 microM, respectively, for pH 4.5 and pH 7.9), hydroxyl radicals (IC50 was 0.11 microM by deoxyribose method and 1.44 microM by electron spin resonance method), and superoxide radicals (IC50 were 2.65 microM) were determined in comparison with positive controls. The inhibitory activities against xanthine oxidase (IC50 were 30.49 microM) were measured. Geraniin also showed dose-dependent inhibitory activities against semicarbazide-sensitive amine oxidase (SSAO, IC50 were 6.58 microM) and against angiotensin converting enzyme (ACE, IC50 were 13.22microM). For kinetic property determinations, geraniin showed competitive inhibitions against SSAO (the apparent inhibition constant, Ki, was 0.70microM) and mixed noncompetitive inhibitions against ACE. Spontaneously hypertensive rats (SHR, 10-week age) were orally administered to once (5 mg geraniin/kg SHR), and changes of systolic blood pressure (SBP) and diastolic blood pressure (DBP) were measured over 24 h and compared with the positive control of captopril (2 mg/kg SHR). The geraniin showed antihypertensive activity in lowering SBP and DBP and showed a significant difference from the blank (distilled water) at 2, 4, 6, 8, and 24 h. Healthy food products could use geraniin for antioxidant protection and therapeutic effects in the future.

Immobilized zinc affinity chromatography of pectin hydroxamic acids for purification of trypsin inhibitors from soybean and sweet potato.

Commercial pectin (with a 94% degree of esterification, DE94) suspended in methanol was reacted with methanolic alkaline hydroxylamine at room temperature for 20 h to prepare pectin hydroxamic acids (PHAs). The prepared PHA was coupled to the epoxy-activated Sepharose 6B gel to get immobilized PHA resins. The immobilized PHA resin was then balanced in column with 2 mM ZnCl2 in 50 mM Tris-HCl buffer (pH 7.9) to test the immobilized Zn-PHA gel as solid phase for immobilized metal affinity chromatography for the purification of trypsin inhibitors (TIs) from soybean and sweet potato. Using TI activity staining, it was found that purified TIs from the commercial soybean and sweet potato after trypsin affinity column purification could be adsorbed onto an immobilized Zn-PHA affinity column and eluted by 100 mM EDTA in 10 mM Tris-HCl buffer (pH 7.9). The immobilized Zn-PHA affinity column was used for TI purifications from crude extracts of sweet potato. The recovery of TI activity for one step was 90%, with 19.74-fold purification increase.