Identification and bioactivity evaluation of ring-opening and lactone forms of enterolactone from sheep fed flaxseed cake.

The previously undescribed lactone ring-opening enterolactone and its sulphate were purified along with the lactone counterparts from the urine of dairy sheep fed flaxseed cake. The structures were determined by NMR and MS analyses. The ring-opening and lactone forms underwent mutual transformation with changes in pH and milk could protect the lactone form. Enterolactone exhibited more effective anti-proliferation activity on MDA-MB-231 breast cancer cells than its ring-opening counterpart, while the ring-opening enterolactone demonstrated more effective anti-osteoporosis activity than the lactone form. The results indicated the potential for targeting biological functions through pH and medium manipulation.

Identification and bioactivity evaluation of flavan-3-ols in the milk of dairy sheep fed Cynomorium songaricum.

Cynomorium songaricum is a traditional medicine and also a food material that is eaten raw or processed as tea or beverages. As a featured plant in semi-desert grasslands, C. songaricum is also eaten by the cattle and sheep in the area. This research study fed dairy sheep C. songaricum to determine the flavan-3-ols in sheep milk. Catechin (Cat), epicatechin (Epi), procyanidin A1 (A1), procyanidin A2 (A2), and procyanidin B1 (B1) were detected in sheep milk with the concentration being Epi > A2 > Cat > B1 > A1 at 24 h after the administration of C. songaricum. Neither A1 nor A2 were detected in the methanol extract of C. songaricum. Cysteine degradation of the plant revealed that in addition to Epi, A2 was the extending unit of the polymeric flavan-3-ols in C. songaricum, indicating that A2 is released digestively from the polymers and enters the milk. Procyanidin B-1 was converted to A1 on incubation in raw but not heated milk, indicating that the A1 in milk is the enzymatically transformed product of B1. Accelerated oxidation showed that the flavan-3-ols, B1, Cat, and Epi significantly protects the unsaturated triacyglycerols in the milk from oxidation. The flavan-3-ol could slow down the oxidation of glutathione and the latter may play an important role in preventing the milk triglycerides from oxidation. Flavan-3-ols are polyphenols with many health benefits. The present research revealed the antioxidant activities of flavan-3-ols that could be absorbed to sheep milk, adding new evidences for the values of these flavan-3-ols and for the milk.

Inhibitory effects and molecular mechanisms of pentagalloyl glucose in combination with 5-FU on aggressive phenotypes of HepG2 cells.

This study examined the inhibition and mechanism of natural product pentagalloyl glucose (PGG) against HepG2 cells and determined the effects of its combination with the clinical chemotherapeutic drug, 5-FU. PGG was found to inhibit the proliferation, migration and invasion of HepG2 cells, induced G1 arrest and apoptosis in both concentration- and time- dependent manners. The combination of PGG and 5-FU had synergistic effects on reversal the aggressive phenotypes of HepG2 cells, increasing the proportion of Bax/Bcl-2, promoting the activation of caspase-9 and caspase-3, and inducing apoptosis. This combination upregulated P27 and cyclin B1, and downregulated cyclin E1, leading to G1 phase arrest. The combination significantly downregulated MDR1 and LRP1, suggesting the potential to reverse the resistance to 5-FU.

Comparison of Antioxidant Constituents of Agriophyllum squarrosum Seed with Conventional Crop Seeds.

Twelve chemical constituents were identified from the Agriophyllum squarrosum seed (ASS). ASS contained large amounts of flavonoids, which were more concentrated in the seed coat. ASS-coat (1 g) contained 335.7 µg flavonoids of rutin equivalent, which was similar to the flavonoid content in soybean (351.2 µg/g), and greater than that in millet, wheat, rice, peanut, and corn. By LC-MS analysis, the major constituents in ASS were 3-O-[α-L-rhamnopyranosyl-(1→6)-ß-D- glucopyranosyl]-7- O-(ß-D-glucopyranosyl)-quercetin (1), rutin (4), quercetin-3-O-ß-D- apiosyl(1→2)-[α-L-rhamnosyl(l→6)]-ß-D-glucoside (2), isorhamnetin-3-O-rutinoside (5), and allantoin (3), compared with isoflavonoids-genistin (16), daidzin (14), and glycitin (18) in soybean. Among constituents in ASS, compounds 1, 2, 4, protocatechuic acid (8), isoquercitrin (11), and luteolin-6-C-glucoside (12) potently scavenged DPPH radicals and intracellular ROS; strongly protected against peroxyl radical-induced DNA scission; and upregulated Nrf2, phosphorylated p38, phosphorylated JNK, and Bcl-2 in HepG2 cells. These results indicate that ASS is rich in antioxidant constituents that can enrich the varieties of food flavonoids, with significant beneficial implications for those who suffer from oxidative stress-related conditions. PRACTICAL APPLICATION: This study found that A. squarrosum seed contains large amounts of antioxidative flavonoids and compared its chemical constituents with those of conventional foods. These results should increase the interest in planting the sand-fixing A. squarrosum on a large scale, thus preventing desertification and providing valuable foods.

Flavan-3-ol-cysteine and acetylcysteine conjugates from edible reagents and the stems of Cynomorium songaricum as potent antioxidants.

Flavan-3-ol derivatives, including 3 cysteine conjugates and 3 acetylcysteine conjugates, were prepared using Cynomorium songaricum and edible reagents. The structures were determined, based on NMR and MS spectra. All compounds had stronger radical-scavenging activity than catechin and epicatechin. Moreover, the cysteine conjugates were active against α-glucosidase, whereas catechin and epicatechin were not. Two acetylcysteine flavan-3-ol conjugates, 4ß-(L-acetylcysteinyl)-epicatechin 3-O-gallate and 4ß-(L-acetylcysteinyl)-epiafzelechin, are novel compounds with better logP values than their cysteine counterparts. These conjugates can be prepared from purified flavan-3-ol polymers or directly from herbal material.

Chemical components and antioxidant activity of the peels of commercial apple-shaped pear (fruit of Pyrus pyrifolia cv. pingguoli).

UNLABELLED: The apple-shaped pear, the fruit of the Pyrus pyrifolia cv. pingguoli (Rosaceae) tree, is one of the most popular fruits in the northern part of China. The current study is the 1st report of its bioactive components. We identified 10 metabolites from the peels (exocarp) of apple-shaped pear and assessed their toxicity. We then compared the anti-oxidant activity, amount of total phenolic compounds, and total condensed tannin content of the peels and flesh (mesocarp) of apple-shaped pear. The 6 major components in the peels and flesh of this fruit were quantified with Ultra Performance Liquid Chromatography-Electrospray Ionization Mass Spectrometry. Results revealed that the peels possessed stronger anti-oxidant activity and contained larger amounts of phenolic compounds than the flesh. These results provide insights into the potential health benefits of this fruit and support the use of the fruit peels and products containing peels or peel components. PRACTICAL APPLICATION: The present research provided evidences that the pulp and peel waste from the juice industry of apple-shaped pear may be a source of useful compounds.

Antioxidant and antiviral activities of lipophilic epigallocatechin gallate (EGCG) derivatives.

The water soluble green tea polyphenol epigallocatechin gallate (EGCG) was lipophilised by esterification with different fatty acids for expanded applications. Four lipophilic ester derivatives of EGCG, namely EGCG-O-tetrastearate, EGCG-O-tetraeicosapentaenoate, EGCG-O-tetradocosahexaenoate, and EGCG-O-octabutyrate, were prepared and evaluated for their antioxidant and antiviral activities in vitro. Incorporation of fatty acids, especially the long chain polyunsaturated fatty acids (PUFA), into EGCG resulted in increased peroxyl radical scavenging activity, as measured by ORAC (oxygen radical absorbance capacity) assay, and metal chelation capacity. However, the esters exhibited decreased reducing power. Antiviral activities of EGCG derivatives were remarkably higher than the parent EGCG molecule, which showed relatively weak effects. The EGCG-PUFA esters were 1700-fold more effective in inhibiting hepatitis C virus (HCV) protease than the positive control embelin. The derivatives also acted as α-glucosidase inhibitors, suggesting their potential in anti-HIV (human immunodeficiency virus) treatment. The results suggest that ester derivatives of EGCG with improved bioactivities may serve as excellent functional food ingredients and natural health products. Moreover, the omega-3 PUFA in the derivatives may also render additional or synergistic health benefits.

Synthesis, anti-HIV and anti-oxidant activities of caffeoyl 5,6-anhydroquinic acid derivatives.

In our continued research on chlorogenic acid analogues and derivatives with improved bioactivity, we have synthesized some caffeoyl 5,6-anhydroquinic acid derivatives. The 1,7 acetonides of chlorogenic acid (15), and of the mono-caffeoyl 5,6-anhydroquinic acids (7-8) showed appreciable anti-HIV activity. The 3,4-dicaffeoyl 5,6-anhydroquinic acid (12) exhibited an anti-HIV activity twice as that of 3,5-dicaffeoylquinic acid (22). The caffeoyl 5,6-anhydroquinic acid derivatives displayed potent anti-oxidant activities. The mono-caffeoyl 5,6-anhydroquinic acids (10-11) were more than twice stronger than chlorogenic acid (21) on SOD-like activity.

Metabolism and pharmacokinetics in rats of ganoderiol F, a highly cytotoxic and antitumor triterpene from Ganoderma lucidum.

The metabolism of ganoderiol F (GF), a cytotoxic and antitumor triterpene from Ganoderma lucidum, by intestinal bacteria and its pharmacokinetics in rats were investigated by using liquid chromatography/mass spectrometry/mass spectrometry (LC/MS/MS). GF was converted to ganodermatriol by anaerobic incubation with bacterial mixtures from rats and humans. This metabolite was detected in rat feces, but not in plasma and urine, after oral administration of GF. The fate of GF after oral (p.o.) and intravenous (i.v.) administration to rats was examined in pharmacokinetics studies. Plasma samples pretreated by solid-phase extraction were quantified by HPLC/MS/MS over a GF concentration range of 1.25-100 ng/ml (S/N = 5). The intra- and interday precision (CV%) was below 8% and accuracy was within the range of 95.9-103.6% for all samples. The range of recovery ratios was 89.2-98.2%. After the administration of GF at 0.5 mg/kg i.v., the plasma concentrations of GF quickly declined and the elimination half-life values (t(1/2alpha) and t(1/2beta)) were about 2.4 and 34.8 min. On the other hand, the elimination half-life values (t(1/2alpha)) after p.o. administration of GF at doses of 20 and 50 mg/kg were 14.4 and 143.3 min for the former, and 18.6 and 114.6 min for the latter. The AUC(0-t) value was 11.17 (ng/ml) h at a GF dose of 0.5 mg/kg i.v., but 49.4 and 111.6 (ng/ml) h at GF doses of 20 and 50 mg/kg p.o., respectively, indicating that the AUC(0-t) value is proportional to the administered oral doses. The estimated absolute bioavailability of GF in rats was F = 0.105.

HCV protease inhibitory, cytotoxic and apoptosis-inducing effects of oleanolic acid derivatives.

PURPOSE: To evaluate oleanolic acid derivatives on liver disease related bioactivities, 29 oleanolic acid derivatives of several series were tested for their inhibitory activity on hepatitis C viral protease and for their cytotoxic effects on Hep G2 cells. RESULTS: The amino derivatives showed potent cytotoxicity, among which, the beta-amino isomer exhibited more distinct cytotoxicity than the alpha-isomer. The cytotoxicity of hemiesters and hemiamides varied as the chain lengths varied. The oxalic and malonic hemiesters showed weaker cytototoxicity than oleanolic acid, while those with longer chain lengths showed higher cytotoxicity. Contrary to the cytotoxic activity, the free amino derivatives showed little inhibitory activity on HCV protease. All the hemiesters and hemiamides showed high activity against HCV protease. CONCLUSION: The findings that addition of amino-group enhanced the cytotoxicity and that introduction of acidic group increased the inhibition on HCV protease may be useful for further design and synthesis of triterpene derivatives as drug candidates for liver diseases.

Inhibitory effects of antrodins A-E from Antrodia cinnamomea and their metabolites on hepatitis C virus protease.

Antrodia cinnamomea is a highly valued folk medicine used for liver cancer, a disease often caused by the long term infection of hepatitis C virus (HCV). In the present study, the maleic and succinic acid constituents (antrodins A-E) of this medicinal fungus, the in vivo metabolites of antrodin C and the analogue of one of the metabolites were tested for their inhibitory activity on HCV protease. Most of the compounds showed potent inhibitory activity, with antrodin A being the most potent (IC(50) = 0.9 microg/mL). Antrodin A was isolated as one of the constituents of A. cinnamomea and was also detected as an in vivo metabolite of the major constituent antrodin C. The mode of inhibition for antrodin A on HCV protease was revealed by a Lineweaver-Burk plot as competitive inhibition. These results strongly support the use of this folk medicine for liver cancer and HCV infection which is a global problem.

Transformation of ergosterol peroxide to cytotoxic substances by rat intestinal bacteria.

Ergosterol peroxide (EPO, 1) is a major antitumor sterol produced by edible or medicinal mushrooms. Following oral administration of 1 to rats or anaerobic in vitro incubation of 1 with rat fecal bacteria, three metabolites were detected and their structures were identified to be 5alpha,6alpha-epoxyergosta-8(14),22-diene-3beta,7alpha-diol (M1, 2), 5alpha,6alpha-epoxyergosta-8,22-diene-3beta,7alpha-diol (M2, 3), and 5alpha,6alpha-epoxy-3beta-hydroxyergosta-22-ene-7-one (M3, 4) by spectroscopic analysis. Of these, M2 and M3 showed more potent inhibitory activity than the original compound 1 against proliferation of CACO-2, WiDr, DLD-1 and Colo320 human colorectal adenocarcinoma cells. These findings suggest that bacterial metabolites of EPO play a significant role in its cytotoxic activity against human colorectal cancer cells.

Estrogenic and anti-estrogenic activities of Cassia tora phenolic constituents.

Through an estrogenic activity bioassay-guided fractionation of the 70% ethanolic extract of Cassia tora seeds two new phenolic triglucosides, torachrysone 8-O-[beta-D-glucopyranosyl(1-->3)-O-beta-D-glucopyranosyl(1-->6)-O-beta-D-glucopyranoside] (1) and toralactone 9-O-[beta-D-glucopyranosyl-(1-->3)-O-beta-D-glucopyranosyl-(1-->6)-O-beta-D-glucopyranoside] (2), along with seven known compounds were isolated. The structures of the new compounds were elucidated on the basis of spectroscopic and chemical evidence. The estrogenic activity of the fractions and the isolated compounds were investigated using the estrogen-dependent proliferation of MCF-7 cells. In addition, the yeast two hybrid assay expressing estrogen receptor alpha (ERalpha) and beta (ERbeta) and the ERalpha competitor screening assay (ligand binding screen) were used to verify the binding affinities of the isolated compounds to ER. Furthermore, a naringinase pre-treatment of the 70% alcoholic extract of Cassia tora seeds resulted in a significant increase in its estrogenic activity. From the naringinase pre-treated extract six compounds were isolated, among which 6-hydroxymusizin and aurantio-obtusin showed the most potent estrogenic activity, while torachrysone, rubrofusarin and toralactone showed a significant anti-estrogenic activity. Finally, the structure requirements responsible for the estrogenic activity of the isolated compounds were studied by investigating the activity of several synthetic compounds and chemically modifying the isolated compounds. The basic nucleus 1,3,8-trihyroxynaphthalene (T(3)HN) was found to play a principal role in the binding affinity of these compounds to ER.

Anti-estrogenic activity of mansorins and mansonones from the heartwood of Mansonia gagei DRUMM.

Through an anti-estrogenic bioassay-guided fractionation of the methanol extract of Mansonia gagei, three new coumarins, called mansorins I (1), II (2) and III (3) and a new naphthoquinone, mansonone I (4), were isolated. Their structures were determined based on their NMR data and CD spectroscopy. The anti-estrogenic activity of the fractions and the isolated compounds were investigated using a yeast two-hybrid assay method expressing estrogen receptors alpha (ERalpha) and beta (ERbeta). In addition, an ERalpha competitor screening system (ligand binding screen) was used to verify the binding affinities of the isolated compounds to the estrogen receptor. 1,2-Naphthoquinones (mansonones) showed more binding affinities to ER in both assay systems. All the tested compounds showed higher binding affinities to ERbeta than to ERalpha in the yeast two-hybrid assay. Mansonones F and S showed the most potent estrogen binding and estrogen antagonistic effects.

The cytotoxic activity of ursolic acid derivatives.

Ursolic acid and 2alpha-hydroxyursolic acid isolated from apple peels were found to show growth inhibitory activity against four tumor cell lines, HL-60, BGC, Bel-7402 and Hela. Structural modifications were performed on the C-3, C-28 and C-11 positions of ursolic acid and the cytotoxicity of the derivatives was evaluated. The SAR revealed that the triterpenes possessing two hydrogen-bond forming groups (an H-donor and a carbonyl group) at positions 3 and 28 exhibit cytotoxic activity. The configuration at C-3 was found to be important for the activity. Introduction of an amino group increased the cytotoxicity greatly. A 3beta-amino derivative was 20 times more potent than the parent ursolic acid. The 28-aminoalkyl dimer compounds showed selective cytotoxicity.

Screening of Chinese and Mongolian herbal drugs for anti-human immunodeficiency virus type 1 (HIV-1) activity.

Water and methanol extracts of 30 Chinese and Mongolian medicinal plants were tested for their human immunodeficiency virus type-1 (HIV-1) inhibitory activity. Of the 60 extracts, 23 showed anti-HIV activity. Bioassay-guided fractionation of one of the most active extracts, the methanol extract of the root tuber of Stephania cepharantha, led to the isolation of two alkaloids, aromoline and FK-3000 as potent inhibitory substances. They completely inhibited the cytopathic effects of HIV-1 on MT-4 cells at 31.3 and 7.8 microg/mL, respectively.