3ß-Palmitoyloxy-olean-12-ene analogs from Sapium lateriflorum (Euphorbiaceae): Their cytotoxic and anti-inflammatory properties and docking studies.

Ten compounds, including a new anti-inflammatory acyl triterpene, 3ß-palmitoyloxy-1ß,11α-dihydroxy-olean-12-ene, were isolated from the bioactive organic extract prepared from the leaves of Sapium lateriflorum (syn: S. nitidum). The isolated compounds were screened for their cytotoxic activity against selected human cancer cell lines and did not display significant activity. They were also evaluated as anti-inflammatory agents in mouse models (TPA-induced edema in the ear and in a carrageenan-induced paw edema model). The results indicated that the new compound, 3ß-palmitoyloxy-1ß,11α-dihydroxy-olean-12-ene, was the compound with major anti-inflammatory activity similar to that of indomethacin, being the hydroxyl at C-11 important for the observed activity. The results of docking studies of the 3ß-palmitoyloxy esters of olean-12-ene with NF-κB and with COX-2 receptors were consistent with possible molecular mechanisms of the anti-inflammatory activity.

Chemical profiling and in-vitro anti-inflammatory activity of bioactive fraction(s) from Trichodesma indicum (L.) R.Br. against LPS induced inflammation in RAW 264.7 murine macrophage cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Trichodesma indicum (L.) R. Br. (family: Boraginaceae) is a medicinal herb largely used to treat arthralgia, rheumatoid arthritis, wound healing, dysentery, etc. It's mechanism of anti-inflammatory activity has not been systematically analyzed yet. AIM OF THE STUDY: The present study was undertaken to examine the anti-inflammatory effects of successive solvent extracts (n-hexane extract (HE), ethyl acetate extract (EA), ethanol extract (EE), aqueous extract (AE) and fractions of HE) from the aerial parts of Trichodesma indicum (TI) against lipopolysaccharide (LPS) stimulated inflammatory reaction using mouse macrophage RAW 264.7 cells. MATERIALS AND METHODS: Cytotoxic effects of the extracts and fractions of TI were assessed by MTT assay. The effect of extracts and fractions on the production of nitric oxide (NO) in RAW 264.7 macrophages were measured using the Griess reagent method. IL - 6, IL - 1ß, TNF-α, iNOS and COX-2 gene expressions were examined by a qRT-PCR method. RESULTS: RAW 264.7 macrophages pretreated with HE, EA, EE and AE of TI showed a significant decrease in the production of proinflammatory cytokines and NO without exhibiting cytotoxicity. The potent HE was fractionated using flash chromatography into FA, FB, FC, FD and FE. Among the five fractions, FE displayed a stronger ability to reduce IL - 1ß, TNF-α, iNOS, COX2 and NO importantly no cytotoxicity was observed. The phytochemical compounds present in FE were further screened by Gas chromatography - Mass spectroscopy (GC-MS). GC-MS analysis revealed that 1,2-benzenedicarboxylic acid diisooctyl ester is the major compound in FE. Molecular docking analysis showed good inhibition of 1,2-benzenedicarboxylic acid diisooctyl ester against TLR-4, NIK and TACE. CONCLUSION: Our results suggested that 1,2-benzenedicarboxylic acid diisooctyl ester could be a potential candidate in alleviating inflammatory reactions in TI.

A Novel Antibacterial Component and the Mechanisms of an Amaranthus tricolor Leaf Ethyl Acetate Extract against Acidovorax avenae subsp. citrulli.

The aim of the present investigation was to determine the active ingredients in Amaranthus tricolor L. leaves and develop a biological pesticide. Organic solvent extraction, column chromatography, liquid chromatography, ODS-C18 reverse elution, Sephadex LH-20 gel filtration, H spectrum, and C spectrum were used to isolate the pure product for an assessment of the agricultural activity and bacteriostatic mechanisms. The results showed that the activity of the crude extract following carbon powder filtration was 1.63-fold that of the non-filtered extract. Further isolation was performed to obtain two pure products, namely, hydroxybenzoic acid (HBA) and benzo[b]furan-2-carboxaldehyde (BFC), and their molecular formulas and molecular weights were C7H6O3 and 138.12, and C9H6O2 and 146.12, respectively. Our study is the first to determine that HBA has bacteriostatic activity (MIC 125 µg/mL) and is also the first to isolate BFC from A. tricolor. The ultrastructure observation results showed that HBA caused the bacteria to become shriveled, distorted, and deformed, as well as exhibit uneven surfaces. After HBA treatment, 70 differentially expressed metabolites were detected in the bacteria, of which 9 were downregulated and 61 were upregulated. The differentially expressed metabolites were mainly strigolactones, organic acids and derivatives, fatty acids, benzene and substituted benzene derivatives, amino acids and associated metabolites, and alcohols and amines. Among all of the downregulated differentially expressed metabolites, MEDP1280 was the most critical, as it participates in many physiological and biochemical processes. The enrichment analysis showed that the differentially expressed metabolites mainly participate in tyrosine metabolism, biosynthesis of amino acids, cysteine and methionine metabolism, and arginine and proline metabolism. Additionally, HBA was found to disrupt cell membrane permeability and integrity, causing the leakage of substances and apoptosis. The physiological and biochemical test results showed that HBA could increase the pyruvate levels in bacteria but could decrease the activities of respiratory enzymes (malate dehydrogenase (MDH) and NADH oxidase) and antioxidant enzymes (superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX)). Inverse molecular docking was used to study the binding between HBA and respiratory and antioxidant enzymes. The results showed that HBA could bind to MDH, NADH oxidase, SOD, and GSH-PX, suggesting that these enzymes may be the effector targets of HBA. Conclusion: The optimal active ingredient in A. tricolor that can inhibit Acidovorax avenae subsp. citrulli was identified as HBA. HBA mainly disrupts the cell membrane, damages the metabolic system, and inhibits respiration and antioxidant enzyme activity to control bacterial growth. These results provide a reference for the further development of biological pesticides.

Separation of Fatty Acid Dimethyl Esters on an Ionic Liquid Gas Chromatographic Column.

An ionic liquid (IL) 111 column was compared with other commonly employed stationary phases including polydimethyl siloxane and polyethylene glycol for the separation of fatty acid monomethyl and dimethyl esters. The fatty acid esters employed in this study were derived from metathesis reactions of vegetable oils both with and without olefins. The IL 111 column demonstrated enhanced performance compared with conventional columns for the separation of these esters. These advantages included significantly enhanced retention of dimethyl esters relative to monomethyl esters, excellent cis/trans isomer separation and the ability to analyze higher carbon number dimethyl esters. As a result, these columns are highly suited for the analysis of mixtures of mono- and dimethyl fatty acid esters found in lipid metathesis reaction products or to determine monofunctional impurities in samples of commercial dimethyl esters.

Preliminary Large Scale Mitigation of 3-Monochloropropane-1, 2-diol (3-MCPD) Esters and Glycidyl Esters in Palm Oil.

Approximately 900 tonne of crude palm oil (CPO) underwent washing using 5 to 10% hot water (90 to 95°C) at a palm oil mill. The aim of the CPO washing was to eliminate and/or reduce total chlorine content present in the conventional CPO, as it is known as the main precursor for the formation of 3-monochloropropane-1, 2-diol esters (3-MCPDE). By a simple hot water washing, more than 85% of the total chlorine was removed. However, washing did not have significant (p > 0.05) effect on other oil quality parameters such as the deterioration of bleachability index (DOBI), free fatty acid (FFA) content and diacylglycerol (DAG) content of the oil. The latter has been established as the main precursor for glycidyl esters (GE) formation. The treated CPO was then transported using tankers and further refined at a commercial refinery. Refining of washed CPO resulted in significantly (p < 0.05) lower formation of 3-MCPDE, but GE content remained slightly high. Post-treatment of refined oil significantly reduced the GE content (p < 0.05) to an acceptable level whilst almost maintaining the low 3-MCPDE level. The study has proven that water washing of CPO prior to refining and subsequent post-refining is so far the most effective way to produce good quality refined oil with considerably low 3-MCPDE and GE contents. Dry fractionation of refined palm oil showed these contaminants partitioned more into the liquid olein fraction compared to the stearin fraction.

Bioactive phenylpropanoid esters of sucrose and anthraquinones from Polygonum cuspidatum.

Four new compounds including two new phenylpropanoid esters of sucrose, polygonusucroside A (1) and B (2); two new anthraquinones, 8-O-ß-d-(6'-galloyl)-glucopyranoside (3) and polyanthraquinoside A (4), together with six known compounds were isolated from Polygonum cuspidatum. Their structures were established using UV, IR, HRESIMS, and NMR data. All compounds were evaluated for their α-glucosidase inhibitory activities and neuroprotective effects. Compounds 5, 7 and 9 showed significant α-glucosidase inhibitory activities with IC50 values of 27.30, 5.51, and 1.09 µmol/L, respectively (acarbose as positive control, IC50 = 6.17 µmol/L). In addition, the assessment of neuroprotective effect showed that compound 3 exhibited remarkable effect against PC12 cells injured by serum-deprivation and compounds 2, 7, and 9 exhibited moderate effects against PC12 cells injured by rotenone.

A new 3,4-seco-lupane triterpenene glycosyl ester from the leaves of Eleutherococcus sessiliflorus.

A new minor 3,4-seco-lupane triterpenene glycosyl ester, named sessiloside-A1 (1), along with three known 3,4-seco-lupane triterpenenes were isolated from the which alcohol extract of the leaves of Eleutherococcus sessiliflorus (Rupr. & Maxim.) S.Y. Hu by silica gel column chromatography, and their structures were determined by spectroscopic methods (UV, IR, NMR and HRMS). Compound 1 was elucidated to be ß-D-glucopyranosyl ester of chiisanogenin. At the same time, a new efficient two-step enzymatic hydrolysis method was established to transform chiisanoside (2) → divaroside (3) → 1.

Antifungal, anti-inflamatory and neuritogenic activity of newly-isolated compounds from Disporopsis aspersa.

A new ester (1) and a terpenoid (2) were isolated from the dried whole plant of Disporopsis aspersa (HUA) ENGL. ex DIELS for the first time and their structures were elucidated, as well as their biological activities are described. The two compounds all showed good antifungal activities, especially furanone (2) exhibited better antifungal activity against Pseudoperonospora cubensis and Phytophthora infestans with EC50 value of 22.82, 18.90 µg/mL, respectively. Compound 1 exhibited a significant promotion on the neurite outgrowth in NGF-induced PC-12 cells, and moderate inhibition on the NO production induced by lipopolysaccharide (LPS) in BV-2 microglial cells.

Oligosaccharide and glucose esters from the roots of Polygala arillata.

The root of plant Polygala arillata has been used in the Oriental medicine as a tonic and for the treatment of certain diseases. Our current research on phytochemical profile of the roots of P. arillata led to the isolation of a new oligosaccharide ester (1, polygaloside), a new glucose ester (7, arillatoside), along with five known sucrose esters (2-6). Their structures were elucidated on the basis of extensive chemical and spectroscopic methods as well as comparison with those reported in the literature. The occurence of various oligosaccharide esters in P. arillata including unique compounds plays taxonomical impact and suggests potential in medicinal uses of the title plant.

Sunflower wax recovered from oil tank settlings: Revaluation of a waste product from the oilseed industry.

BACKGROUND: The sunflower oil industry produces a large amount of waste that is not currently commercially exploited, as in the case of oil-tank settlings. The recovery of a high value-added by-product, such as sunflower wax, would increase the commercial value of this waste. In this original research paper, a method that allows the recovery and purification of this by-product was developed. The wax was characterized and its potential use as an organogelator agent was investigated. RESULTS: The waste sample was composed of 45.1% oily material, 16.9% of this being impure waxes. Purification was performed through two different methods, obtaining three waxes with different degrees of purity. All the waxes were composed of wax esters with a range of 40-60 carbon atoms, exhibiting traces of carotenes, free fatty acids, and free fatty alcohols. The presence of phospholipids was observed in two of them. The third wax presented a higher total wax ester content and physicochemical characteristics (color and thermal behavior) similar to those of commercial sunflower waxes, and was the most efficient organogelator agent, requiring only a small amount of wax (1.5%) to structure high oleic sunflower oil. CONCLUSION: It was verified that sunflower wax could be recovered from oil-tank settlings. A purification method that allowed sunflower wax with similar physicochemical properties to those of commercial waxes to be obtained was also developed. The purified waxes were capable of structuring high oleic sunflower oil. © 2019 Society of Chemical Industry.

In Silico, Cytotoxic and Antioxidant Potential of Novel Ester, 3-hydroxyoctyl -5- trans-docosenoate Isolated from Anchusa arvensis (L.) M.Bieb. Against HepG-2 Cancer Cells.

BACKGROUND: Cancer is one of the chronic health conditions worldwide. Various therapeutically active compounds from medicinal plants were the current focus of this research in order to uncover a treatment regimen for cancer. Anchusa arvensis (A. anchusa) (L.) M.Bieb. contains many biologically active compounds. METHODS: In the current study, new ester 3-hydroxyoctyl -5- trans-docosenoate (compound-1) was isolated from the chloroform soluble fraction of A. anchusa using column chromatography. Using MTT assay, the anticancer effect of the compound was determined in human hepatocellular carcinoma cells (HepG-2) compared with normal epithelial cell line (Vero). DPPH and ABTS radical scavenging assays were performed to assess the antioxidant potential. The Molecular Operating Environment (MOE-2016) tool was used against tyrosine kinase. RESULTS: The structure of the compound was elucidated based on IR, EI, and NMR spectroscopy technique. It exhibited a considerable cytotoxic effect against HepG-2 cell lines with IC50 value of 6.50 ± 0.70 µg/mL in comparison to positive control (doxorubicin) which showed IC50 value of 1.3±0.21 µg/mL. The compound did not show a cytotoxic effect against normal epithelial cell line (Vero). The compound also exhibited significant DPHH scavenging ability with IC50 value of 12 ± 0.80 µg/mL, whereas ascorbic acid, used as positive control, demonstrated activity with IC50 = 05 ± 0.15 µg/mL. Similarly, it showed ABTS radical scavenging ability (IC50 = 130 ± 0.20 µg/mL) compared with the value obtained for ascorbic acid (06 ± 0.85 µg/mL). In docking studies using MOE-2016 tool, it was observed that compound-1 was highly bound to tyrosine kinase by having two hydrogen bonds at the hinge region. This good bonding network by the compound might be one of the reasons for showing significant activity against this enzyme. CONCLUSION: Our findings led to the isolation of a new compound from A. anchusa which has significant cytotoxic activity against HepG-2 cell lines with marked antioxidant potential.

Montbresides A-D: antibacterial p-coumaroyl esters of a new sucrose-based tetrasaccharide from Crocosmia × crocosmiiflora (montbretia) flowers.

Crocosmia × crocosmiiflora (montbretia) flowers yielded four esters (montbresides A-D) of a new sucrose-based tetrasaccharide, 3-O-ß-d-glucopyranosyl-4´-O-α-d-rhamnopyranosyl-sucrose [ß-d-Glc-(1 → 3)-α-d-Glc-(1↔2)-ß-d-Fru-(4 ← 1)-α-d-Rha]. All four possess O-p-coumaroyl residues on C-3 of fructose and C-4 of α-glucose, plus O-acetyl residues on C-2 and C-3 of rhamnose and C-6 of fructose. Montbresides A and B are additionally O-acetylated on C-1 of fructose. The p-coumaroyls are trans- in montbresides A and C and cis- in B and D. Elemental compositions were determined from MS data, and structures from 1D and 2D NMR spectra. Monosaccharide residues were identified from selective 1D TOCSY spectra and TLC, and acylation sites from 2D HMBC spectra. Enantiomers were distinguished by enzymic digestion. Montbretia flower extracts were cytotoxic against six human cancerous cell-lines, but purified montbresides lacked cytotoxicity. Each montbreside displayed antibacterial activity against Staphylococcus aureus (minimal inhibitory concentration ~6 µg/ml). Montbretia is a potential source of new cytotoxins and antibacterial agents.

Novel sodium alkyl-1,3-disulfates, anionic biosurfactants produced from microbial polyesters.

A sodium alkyl disulfate mixture (SADM) synthesised from microbially produced 3-hydroxy fatty acids methyl esters (HFAMEs), showed 13-fold surface tension decrease when compared with the reference surfactant sodium dodecyl sulfate (SDS). Polyhydroxyalkanoates, accumulated by bacteria intracellularly when supplied with a mixture of fatty acids derived from hydrolysed rapeseed oil, were isolated, depolymerised and methylated to produce HFAMEs in very high yield (90%). A sequential chemical reduction and sulfation of the HFAMEs produced the sodium alkyl disulfates in high yields (>65%). SADM performs also 1.3-times better than dodecyl (1,3) disulfate, in surface tension tests. SADM shows also the formation of a specific critical micelle concentration (CMC) at a concentration 21-fold lower than SDS. The wettability of the SADM mixture is similar to SDS but the foaming volume of SADM is 1.5-fold higher. The foam is also more stable with its volume decreasing 3 times slower over time compared to SDS at their respective CMC values. Established sulfation technologies in chemical manufacturing could use the 3-hydroxy fatty acids methyl esters moiety (3-HFAME) given its origin from rapeseed oil and the extra OH residue on 3-position in the molecule, which affords the opportunity to produce disulfate surfactants with a proven superior performance to monosulphated surfactants. Thus, not only addressing environmental issues by avoiding threats of deforestation and monocultivation associated with palm oil use but also achieve a higher performance with lower use of surfactants.

Phenolic cyclobutantetraol esters from Scindapsus officinalis (Roxb.) Schott.

Five new caffeoyl cyclobutantetraol esters (1-5) and one hydroxycinnamoyl cyclobutantetraol ester (6), were isolated from Scindapsus officinalis (Roxb.) Schott, which were the first reported phenolic metabolites incorporating a cyclobutantetraol in nature. Structures of the isolated compounds, including absolute configurations, were elucidated by spectroscopic analysis, especially 2D NMR techniques and exciton chirality CD (ECCD) method. All isolates were evaluated for cytotoxic activity toward MCF-7 human breast cancer cell, anti-inflammatory activity against nitric oxide (NO) production, and their antioxidative activity in the 1,1-diphenyl-2-picrylhydrazyl assay in vitro. Compound 1 showed strong antioxidative activity with IC50 value of 59.2 µM, and compounds 1-6 exhibited weak inhibitory effects on NO production, while hardly showing any cytotoxic effects against MCF-7 cell.

Removal of phthalic acid esters from sea buckthorn (Hippophae rhamnoides L.) pulp oil by steam distillation and molecular distillation.

Sea buckthorn (Hippophae rhamnoides L.) pulp oils (SPOs) are rich in a variety of beneficial bioactive ingredients. Nevertheless, SPOs would be exposed to plastic equipment during processing, resulted in increasing phthalates contents and edible risk, as well as affecting oil quality. For these reasons, the effects of two stages steam distillation (SD2) and two stages molecular distillation (MD2) on phthalic acid esters (PAEs) content were investigated and compared in the present work. Compared with SD2, MD2 showed higher removal rates of seven selected PAEs from the SPO. Even if the initial concentration of DBP and DEHP in R-SPO were 1.626 and 10.933 mg/kg respectively, the concentration of DBP and DINP could be reduced below the limit set by China government after treated with MD2. Besides that, there was no trans-fatty acids generated in SPO during the distillation process.

Marigold carotenoids: Much more than lutein esters.

Carotenoids constitute a large group of lipophilic pigments whose health-promoting benefits have been widely recognized. Hydroxy-containing carotenoids can be found in both free form or esterified with fatty acids in several plant matrices, but the native carotenoid profile is overall poorly explored due to the difficulty of analyzing carotenoid esters. One of the main natural sources of carotenoids is the marigold flower, which has been extensively used by the industry for the production of food colorants or supplements, both often manufactured with no saponification process. Although lutein esters are well established as the major compounds naturally found in marigold petals and their products, carotenoid esters other than the lutein ones have not been extensively examined. We carried out a comprehensive identification of carotenoids and carotenoid esters from marigold petals by LC-DAD-(APCI+)MS/MS. Whereas 18 carotenoids were identified in the saponified extract, 56 were identified when no saponification procedure was carried out: 6 free carotenoids, 20 monoesters and 30 diesters. This is the first time that esters of zeaxanthin, violaxanthin, auroxanthin, zeinoxanthin and ß-cryptoxanthin are identified in marigold. The structural information obtained through characteristic fragmentation patterns and diagnostic fragments in MS and MS/MS spectra (APCI+) sustained the differentiation between carotenoid esters with similar characteristics. Therefore, the separation of carotenoids by reversed-phase liquid chromatography using C30 columns in combination with DAD and APCI-MS/MS detection allowed high sensitivity and selectivity for carotenoid ester analysis.

Washing bleached palm oil to reduce monochloropropanediols and glycidyl esters.

The formation of toxic compounds, potentially carcinogenic, during food processing has been considered an important food safety issue. Among them, particular attention has been given to 3-monochloropropane-1,2-diol esters (3-MCPDE), 2-monochloropropane-1,3-diol esters (2-MCPDE) and glycidyl esters (GE), which can be formed during vegetable oil refining, especially palm oil. These substances may pose a health risk to humans due to their toxicity and carcinogenicity. The aim of this study was to investigate the effect of washing bleached palm oil (BPO) with different solvents, and evaluate the reduction of 3-MCPDE, 2-MCPDE and GE as well as assess the quality parameters of the final product. For this purpose, we used two types of washing with different solvents. A single washing was carried out in one step and a double washing in two steps using a solvent gradient. Single washing had a limited reduction in the levels of 3-MCPDE and 2-MCPDE and resulted in an increased level of GE, whereas double washing slightly reduced 3-MCPDE and 2-MCPDE and resulted in a significant reduction of GE levels. The reduction achieved in this study was up to 17.1% for 3-MCPDE, 56.4% for 2-MCPDE and 76.9% for GE levels. The reduction of 3-MCPDE and 2-MCPDE might be due to the removal of part of the ethanol-soluble chlorinated precursors from the oil which suggests that highly lipophilic forms of these substances are present in BPO. The substantial reduction on GE levels might be associated with the removal of the precursors present in the oil such as diacylglycerols. Thus, the washing treatment could be used as a supplementary strategy to reduce processing contaminants from palm oil, especially GEs.

Effect of Skin Maceration Treatment on Aroma Profiles of Kiwi Wines Elaborated with Actinidia deliciosa "Xuxiang" and A. chinensis "Hort16A".

Background: Kiwi has become an important and promising fruit to ferment kiwi wine in recent years in China. Objective: The objective of this study was to evaluate the effect of skin maceration treatment on aroma profiles of kiwi wine elaborated with two representative kiwi varieties (Actinidia deliciosa "Xuxiang" and A. chinensis "Hort16A"). Methods: Aroma profiles were characterized using solid-phase microextraction GC-MS method. Principal component analysis was used to separate and group the wines as well as for identifying the aroma components that best differentiate the wines. Results: Esters and alcohols were the two most abundant compounds in kiwi wine. Skin maceration treatment gave rise to a positive effect of aroma profiles, resulting in a significant increase of terpenes. A total of 11 volatile compounds were found at concentrations higher than their odor threshold in kiwi wine samples. Conclusions: The study could play a role in laying a foundation for the development of the kiwi fruit wine industry. Highlights: Different aroma profiles were presented because of variety differences. Skin maceration treatment gave rise to a positive effect of aroma profiles, which resulted in a significant increase of terpenes.

Isolation of a new flavonoid and waste to wealth recovery of 6-O-Ascorbyl Esters from Seeds of Aegle marmelos (family- Rutaceae).

Aegle marmelos is a plant species native to India. Commercially available food products such as jam, jelly, candy, squash etc. are prepared from ripe fruit pulp of A. marmelos. Ripe fruit processing accounts for 60% of whole fruit mass while 40% remains unutilized and generates waste (hard shell, pomace, fiber and seeds) on a massive scale which do not have high value applications. A new flavone 3,5,7-trihydroxy-2-(4'-hydroxy-3'-isopentyloxyphenyl-4H-chromen-4-one (5) in addition to the known compounds 1-4, has been isolated from seeds of A. marmelos. Also, compound 7-(3'-methylbut-2'-enyloxy)-2H-chromen-2-one (2) has been isolated for the first time from A. marmelos. The structure of compounds 1-5 was determined by spectral analysis (UV, IR, NMR, etc.). Additionally, the non-edible oil obtained from seeds was investigated for waste to wealth recovery of 6-O-ascorbyl esters in high regioselectivity via one step semi-synthetic approach in the presence of ascorbic acid and H2SO4 at ambient temperature.

Isolation and Purification of Galloyl, Caffeoyl, and Hexahydroxydiphenoyl Esters of Glucoses from Balanophora simaoensis by High-Speed Countercurrent Chromatography and Their Antioxidant Activities In Vitro.

High-speed counter-current chromatography was used to separate and purify galloyl, caffeoyl, and hexahydroxydiphenoyl esters of glucoses from the aerial parts of the parasitic plant Balanophora simaoensis for the first time using n-hexane-ethyl acetate-methanol-water (1:2:1:2, v/v) as the optimum solvent system. Accordingly, 1-O-(E)-caffeoyl-3-O-galloyl-ß-d-glucopyranose (I, 12.5 mg), 1-O-(E)-caffeoyl-3-O-galloyl-4,6-(S)-hexahydroxydiphenoyl-ß-d-glucopyranose (II, 27.2 mg), and 1-O-(E)-caffeoyl-4,6-(S)-hexahydroxydiphenoyl-ß-d-glucopyranose (III, 52.8 mg) with 98.0%, 98.5%, and 98.7% purities, respectively, were purified from 210 mg crude extract of B. simaoensis in a one-step separation. The structures of the glucose esters were identified by electrospray ionization mass spectrometry (ESI-MS) and nuclear magnetic resonance spectra (NMR). Their antioxidant activities were evaluated by measuring their inhibition activity on liver microsomal lipid peroxidation induced by the Fe2+-Cys system in vitro. Compounds I⁻III showed significant antioxidant activities with IC50 values ranging from 2.51 to 6.68 µm, respectively.