Discovery of natural AMPK activator from the fruits of Xanthium sibiricum Patr.: Xanthiumine A, protoberberine alkaloid with unique C28 skeleton.

Two protoberberine alkaloids with a unique C28 skeleton, named xanthiumines A (1) and B (2), respectively, were isolated from the fruits of Xanthium sibiricum Patr. Their structures including absolute configurations were unequivocally established by the comprehensive NMR and MS spectroscopic data analysis together with gauge-independent atomic orbital (GIAO) NMR calculations, and electronic circular dichroism (ECD) calculations. Compounds 1 and 2 are the first examples of natural protoberberine alkaloid with a phenolic acid group at C-13a. Their plausible biosynthetic pathway was proposed on the basis of the coexisting alkaloid monomer as the precursor. Furthermore, the effects and related molecular mechanism of compound 1 on hepatic lipid accumulation were also investigated in oleic acid (OA)-treated HepG2 cells.

Unprecedented diterpenoid dimers with soluble epoxide hydrolase inhibitory effect from Euphorbia fischeriana.

Biseuphoids A (1) and B (2), two unprecedented ent-abietane-type diterpenoid dimers linked by monomeric blocks through C-17-C-12' and C-17-C-11', respectively, were isolated from Euphorbia fischeriana, along with their biogenesis related diterpenoid monomers, 17-hydroxyjolkinolide B (3), caudicifolin (4), and fischeriabietane C (5). Their structures were elucidated by extensive spectroscopy assisted by quantum chemical NMR and ECD calculations. The unusual dimeric skeletons are possibly derived from the adduct of diterpenoid monomers through Michael-like reactions. The novel dimers 1 and 2 exhibited inhibitory activities on soluble epoxide hydrolase (sEH) with IC50 values of 8.17 and 5.61 µM, respectively. Molecular dynamics studies illustrated that both 1 and 2 can occupy the catalytic pocket of sEH by forming stable hydrogen bonds with the key amino acid residues including Gln384, Asn378, Pro361, Ala365, Asn366, and Asn472.

Protostane-type triterpenoids as natural soluble epoxide hydrolase inhibitors: Inhibition potentials and molecular dynamics.

The inhibition of soluble epoxide hydrolase (sEH) is a promising therapeutic approach to treat inflammation and other disorders. In our present investigation on searching for sEH inhibitors from traditional Chinese medicines, we found that Alisma orientale displayed inhibition of sEH. We constructed a small library of protostane-type triterpenoids (1-25) isolated from A. orientale, and screened their inhibitory activities. Alismanin B (1), 11-deoxy-25-anhydro alisol E (4), 11-deoxy alisol B (5), and 25-O-ethyl alisol A (15) displayed concentration-dependently inhibitory activities against sEH with IC50 values from 3.40 ± 0.57 µM to 9.57 ± 0.88 µM. 11-Deoxy-25-anhydro alisol E (4) and 11-deoxy alisol B (5) were defined as mixed-type competitive inhibitors with Ki values of 12.6 and 3.48 µM, respectively, based on the result of inhibition kinetics. The potential interaction mechanism of 11-deoxy alisol B (5) with sEH was analyzed by molecular docking and molecular dynamics, revealing that amino acid residues Trp336 and Tyr466 were vital for its inhibitory activity.

Berberrubine and its analog, hydroxypropyl-berberrubine, regulate LDLR and PCSK9 expression via the ERK signal pathway to exert cholesterol-lowering effects in human hepatoma HepG2 cells.

Berberine (BBR), the major isoquinoline alkaloid in Chinese herb Rhizoma coptidis, has significant lipid-lowering effect by upregulating hepatic low-density lipoprotein receptor (LDLR) expression. In a previous study, we have indicated that berberrubine (M3), a major metabolite of BBR in vivo, displays the most potential hypolipidemic effects via upregulating LDLR expression in human hepatoma (HepG2) cells compared with BBR and 3 other metabolites. Accordingly, 9 M3 analogs (A1-A9) were modified at the C9 position. We aimed to find a new promising agent by evaluating the cholesterol-lowering effect and clarifying the related molecular mechanism. In the current study, the cellular cholesterol content was assayed with a commercial cholesterol assay kit. Real-time polymerase chain reaction and Western blot assay were used to explore the molecular mechanism of M3 and its analogs on the hypolipidemic effect. Among M3 and its analogs, hydroxypropyl-berberrubine (A8) exhibited the highest potential effects on the upregulation of LDLR expression, which was accompanied by a steady decline of proprotein convertase subtilisin/kexin type 9 (PCSK9) messenger RNA and protein levels. Furthermore, inhibition of extracellular signal-regulated kinase (ERK) activity with PD98059 prevented the upregulation of LDLR and downregulation of PCSK9 induced by A8. The current study revealed that M3 and its structurally modified analog, A8, could regulate hepatic LDLR and PCSK9 expression to exert lipid-lowering effects via the ERK signal pathway, while A8 showed a stronger effect and might be a promising drug candidate against hyperlipidemia.

Two pairs of phenylpropanoid enantiomers from the leaves of Eucommia ulmoides.

Two pairs of phenylpropanoid enantiomers, (+)-(7S,8S)-alatusol D (1a), (-)-(7R,8R)-alatusol D (1b), (-)-(7S,8R)-alatusol D (2a) and (+)-(7R,8S)-alatusol D (2b) were isolated from the leaves of Eucommia ulmoides Oliver. Among them, 1a and 2b were firstly obtained by chiral enantiomeric resolution. Their structures were elucidated based on extensive spectroscopic analysis and the induced CD (ICD) spectrum caused by adding Mo2(AcO)4 in DMSO. All compounds were tested on Hep G2 tumor cell lines. However, none of the compounds showed potential cytotoxic activity against Hep G2 in vitro.

A high-resolution α-glucosidase inhibition profiling for targeted identification of natural antidiabetic products from Lycopodiella cernua (L.) Pic. Serm and their inhibitory mechanism study.

The targeted identification of α-glucosidase inhibitors from the crude ethyl acetate of Lycopodiella cernua (L.) Pic. Serm (L.cernua) was guided by high-resolution inhibition profiling. The α-glucosidase inhibition profiling and HPLC-QTOF-MS showed tannins and serratenes were the corresponding antidiabetic constituents. Two new serratenes named 3ß, 21ß-dihydroxyserra-14-en-24-oic acid-3ß-(4'-methoxy-5'-hydroxybenzoate) (4), 3ß, 21α-dihydroxyserra-14-en-24-oic acid-3ß-(4'-methoxy-5'-hydroxybenzoate) (7), together with two known compounds (5 and 6) were isolated. Their structures were elucidated by HR-ESI-MS and NMR. Compounds 5-7 inhibited the α-glucosidase activity in a non-competitive manner with Ki values ranging from 1.29 to 12.9 µM. The molecular docking result unveiled that 4-7 bound to the residues at the channel site, which enabled to block the substrate access. In addition, the molecular dynamics (MD) simulation of the most active compound 7 and α-glucosidase indicated the 4'-methoxy-5'-hydroxybenzoate group formed the stable hydrogen bonds and pi-pi T-shaped interactions with Arg312, Gln350 and Phe300 residues, while the rings D and E were stabilized by hydrophobic interaction.

Inhibition effect of 1-acetoxy-6α-(2-methylbutyryl)eriolanolide toward soluble epoxide hydrolase: Multispectral analysis, molecular dynamics simulation, biochemical, and in vitro cell-based studies.

Soluble epoxide hydrolase (sEH) serves as a potential target in inflammation-related diseases. Based on the bioactivity-guided separation, a new sesquiterpenoid inulajaponoid A (1) was isolated from Inula japonica with a sEH inhibitory effect, together with five known compounds, such as 1-O-acetyl-6-O-isobutyrylbritannilactone (2), 6ß-hydroxytomentosin (3), 1ß,8ß-dihydroxyeudesma-4(15),11(13)-dien-12,6α-olide (4), (4S,6S,7S,8R)-1-O-acetyl-6-O-(3-methylvaleryloxy)-britannilactone (5), and 1-acetoxy-6α-(2-methylbutyryl)eriolanolide (6). Among them, compounds 1 and 6 were assigned as mixed and uncompetitive inhibitors, respectively. The result of immunoprecipitation (IP)-MS demonstrated the specific binding of compound 6 to sEH in the complex system, which was further confirmed by the fluorescence-based binding assay showing its equilibrium dissociation constant (Kd = 2.43 µM). The detail molecular stimulation revealed the mechanism of action of compound 6 with sEH through the hydrogen bond of amino acid residue Gln384. Furthermore, this natural sEH inhibitor (6) could suppress the MAPK/NF-κB activation to regulate inflammatory mediators, such as NO, TNF-α, and IL-6, which confirmed the anti-inflammatory effect of inhibition of sEH by 6. These findings provided a useful insight to develop sEH inhibitors upon the sesquiterpenoids.

Room Temperature Engineering Crystal Facet of Cu2O for Photocatalytic Degradation of Methyl Orange.

Cuprous oxide (Cu2O) has received enormous interest for photocatalysis owing to its narrow band gap of 2.17 eV, which is beneficial for visible-light absorption. In this work, we succeeded in synthesizing Cu2O nanocrystals with two morphologies, cube and sphere, at room temperature via a simple wet-chemistry strategy. The morphologies of Cu2O change from cube to sphere when adding PVP from 0 g to 4 g and the mainly exposed crystal faces of cubic and spherical Cu2O are (100) and (111), respectively. The photocatalytic properties of the as-prepared Cu2O were evaluated by the photocatalytic degradation of methyl orange (MO). Cubic Cu2O(100) showed excellent photocatalytic activity. After the optical and photoelectric properties were investigated, we found that cubic Cu2O(100) has better photoelectric separation efficiency than spherical Cu2O(111). Finally, the possible mechanism was proposed for cubic Cu2O(100) degrading MO under visible light.

SILAC-based chemoproteomics reveals a neoligan analogue as an anti-inflammatory agent targeting IRGM to ameliorate cytokine storm.

Cytokine storm is a key feature of sepsis and severe stage of COVID-19, and the immunosuppression after excessive immune activation is a substantial hazard to human life. Both pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs) are recognized by various pattern recognition receptors (PRRs), which lead to the immune response. A number of neolignan analogues were synthesized in this work and showed powerful anti-inflammation properties linked to the response to innate and adaptive immunity, as well as NP-7 showed considerable anti-inflammatory activity at 100 nM. On the sepsis model caused by cecum ligation and puncture (CLP) in C57BL/6J mice, NP-7 displayed a strong regulatory influence on cytokine release. Then a photo-affinity probe of NP-7 was synthesized and chemoproteomics based on stable isotope labeling with amino acids in cell cultures (SILAC) identified Immunity-related GTPase M (IRGM) as a target suppressing cytokine storm, which was verified by competitive pull-down, cellular thermal shift assay (CETSA), drug affinity responsive target stability (DARTS) and molecular dynamics simulations.

Biotransformation of 18ß-Glycyrrhetinic Acid by Human Intestinal Fungus Aspergillus niger RG13B1 and the Potential Anti-Inflammatory Mechanism of Its Metabolites.

18ß-Glycyrrhetinic acid (GA) is a triterpenoid possessing an anti-inflammatory activity in vivo, while the low bioavailability limits its application due to its intestinal accumulation. In order to investigate the metabolism of GA in intestinal microbes, it was incubated with human intestinal fungus Aspergillus niger RG13B1, finally leading to the isolation and identification of three new metabolites (1-3) and three known metabolites (4-6) based on 1D and 2D NMR and high-resolution electrospray ionization mass spectroscopy spectra. Metabolite 6 could target myeloid differentiation protein 2 (MD2) to suppress the activation of nuclear factor-kappa B (NF-κB) signaling pathway via inhibiting the nuclear translocation of p65 to downregulate its target proteins and genes in lipopolysaccharide (LPS)-mediated RAW264.7 cells. Molecular dynamics suggested that metabolite 6 interacted with MD2 through the hydrogen bond of amino acid residue Arg90. These findings demonstrated that metabolite 6 could serve as a potential candidate to develop the new inhibitors of MD2.

Amentoflavone from Selaginella tamariscina as a potent inhibitor of gut bacterial ß-glucuronidase: Inhibition kinetics and molecular dynamics stimulation.

Gut bacterial ß-glucuronidase (GUS) plays a pivotal role in the metabolism and reactivation of a vast of glucuronide conjugates of both endogenous and xenobiotic compounds in the gastrointestinal tract of human, which has been implicated in certain drug-induced gastrointestinal tract (GI) toxicity in clinic. Inhibitors of gut microbial GUS exhibited great potentials in relieving the drug-induced GI toxicity. In this study, Selaginella tamariscina and its major biflavonoid amentoflavone (AMF) were evaluated for their inhibitory activity against Escherichia coli GUS. Two selective probe substrates for GUS (a specific fluorescent probe substrate for GUS, DDAOG and a classical drug substrate for GUS, SN38G) were used in parallel for charactering the inhibition behaviors. Both the extract of S. tamariscina and its major biflavonoid AMF displayed evident inhibitory effects on GUS, and the IC50 values of AMF against GUS mediated DDAOG and SN-38G hydrolysis were 0.62 and 0.49 µM, respectively. Inhibition kinetics studies indicated that AMF showed mixed type inhibition for GUS-mediated DDAOG hydrolysis, while displayed competitive type inhibition against GUS-mediated SN-38G hydrolysis, with the Ki values of 0.24 and 1.25 µM, respectively. Molecular docking studies and molecular dynamics stimulation results clarified the role of amino acid residues Leu361, Ile363, and Glu413 in the inhibition of AMF on GUS. These results provided some foundations for the potential clinical utility of S. tamariscina and its major biflavonoid AMF for treating drug-induced enteropathy.

Investigation of the inhibitory effect of protostanes on human carboxylesterase 2 and their interaction: Inhibition kinetics and molecular stimulations.

Carboxylesterase 2 (CES 2), plays a pivotal role in endobiotic homeostasis and xenobiotic metabolism. Protostanes, the major constituents of the genus Alisma, display a series of pharmacological activities. Despite the extensive studies of pharmacological activities, the investigation on inhibitory effects of protostanes against CES 2 is rarely reported. In this study, the inhibitory activities of a library of protostanes (1-25) against human CES 2 were investigated for the first time, using 6,8-dichloro-9,9-dimethyl-7-oxo-7,9-dihydroacridin-2-yl benzoate (DDAB) as the specific fluorescent probe for human CES 2. Compounds 1, 2, 7, 8, 12, 13, 18, 19, and 25 showed strong inhibitory effects towards CES 2. For the most potent compounds 1, 7, 13, and 25, the inhibition kinetics were further investigated, and these four protostanes were all uncompetitive inhibitors against human CES 2 with the inhibition constant (Ki) values ranging from 0.89 µM to 2.83 µM. In addition, molecular docking and molecular dynamics stimulation were employed to analyze the potential interactions between these protostanes and CES 2, and amino acid residue Gln422 was identified to play a crucial role in the strong inhibition of protostanes towards CES 2.

An integrated approach based on phytochemistry, network pharmacology and metabolomics reveals the mechanism of action of Xanthium strumarium L. for allergic rhinitis.

Xanthium strumarium L. (XS) is a traditional Chinese medicine (TCM) that has been widely used in Chinese medicine prescription for allergic rhinitis (AR). However, the action mechanisms of XS on the therapeutic effects on AR remain elusive. Herein, an integrated approach of phytochemistry, network pharmacology and metabolomics was first applied to uncover the action mechanisms of XS for AR. The therapeutic effect of XS extract on AR was evaluated in rat models of ovalbumin (OVA)-induced AR. The cytokine levels in rat serum and histopathological changes of nasal mucosa were assessed after oral treatment with XS. Chemical compositions of XS were elucidated by phytochemical methods, and active ingredients were identified via ADME-TOX screening in silico. Network pharmacology was performed to establish and analyze the compound-target-disease network so as to find the possible mechanism of XS in treating AR. In addition, metabolomics analysis was applied to investigate the changes in the endogenous metabolite levels that result from XS treatments. As result, the XS extract significantly increased the serum concentrations of IL-2 and reduced the levels of serum IL-4, while XS could ameliorate inflammation in the nasal sub-mucosal area, indicating that XS has significant therapeutic effects on AR model rats. Furthermore, a total of 119 compounds were isolated from XS, and 59 of these compounds were identified as active ingredients through ADME-TOX screening in silico. An in-depth analysis of the network pharmacology implied that the active ingredients of XS could regulate the inflammatory response via "multi-component, multi-target" patterns. In combination with the results of metabolomics, we found that the active ingredients of XS have a beneficial effect on AR through regulating the metabolism of arachidonic acid, which was reflected by medicating the Fc epsilon RI signaling pathway, and the neuroactive ligand-receptor interaction pathway, as well as the key proteins in arachidonic acid metabolism, such as PTGS2, PTGS1, PTGES and ALOX5. Additionally, molecular docking showed that multiple compounds have better binding with PTGS2 and ALOX5, which might be two crucial targets. Overall, these results suggest that the treatment of XS for AR is realized by regulating the metabolism of arachidonic acid via a combination form. This study provides the basis for clinical applications of XS.

The inhibition effect of uncarialin A on voltage-dependent L-type calcium channel subunit alpha-1C: Inhibition potential and molecular stimulation.

Cardiovascular diseases, such as hypertension and cardiac failure, have become the most major and global cause for threatening human health in recent years. Uncaria rhynchophylla as a traditional Chinese medicine is widely used to treat hypertension for a long history, whereas its medicinal effective components and potential action mechanism are uncertain. Therefore, twenty-four alkaloids (1-24) isolated from U. rhynchophylla were assayed for their relaxant effects against phenylephrine (Phe)-induced contraction of rat mesenteric arteries. Among them, we surprisingly found that uncarialin A (21) exhibited most potent relaxation effect against Phe-induced contraction (IC50 = 0.18 µM) in the manner of independent on endothelium-derived vasorelaxing factors and endothelium. All the experiments including measurement of Ca2+ in vascular smooth muscle cells (VSMCs) by fluorescence microscopy, whole-cell path clamp, molecular docking, and molecular dynamics, demonstrated that uncarialin A (21) could significantly inhibit L-type calcium channel subunit alpha-1C (Cav1.2) via the hydrogen bond interaction with amino acid residue Met1186, allowing the inhibition of Ca2+ inward current. Our results suggested that uncarialin A (21) could be served as a potential L-type Cav1.2 blocker in the effective treatment of cardiovascular diseases.

The study of inhibitory effect of natural flavonoids toward ß-glucuronidase and interaction of flavonoids with ß-glucuronidase.

ß-Glucuronidase plays a vital role in the metabolism of drugs and endogenous substance. Herein, we assayed the inhibitory effects of thirty-six flavonoids (1-36) toward ß-glucuronidase (Escherichia coli) using the probe reaction of DDAO-glu hydrolysis. The results showed that kushenol X (6), (2S)-farrerol (10), 5,7,2'-trihydroxy-8,6'-dimethoxy flavone (20), demethylbellidifolin (31), and gentisin (32) exhibited potent inhibitory activities toward ß-glucuronidase with the IC50 values of 2.07 ± 0.26, 8.95 ± 0.74, 4.97 ± 0.61, 0.91 ± 0.11, and 0.68 ± 0.10 µM, respectively. Furthermore, the inhibition kinetics studies indicated that demethylbellidifolin (31) and gentisin (32) exhibited mixed-type inhibiton toward ß-glucuronidase, the Ki values were caculated to be 4.05 and 2.02 µM, respectively. Additionally, the circular change of dichroism (CD) spectrum verified the interaction between demethylbellidifolin (31) and gentisin (32) with ß-glucuronidase; following by the molecular docking and molecular dynamics further revealed the potential interaction amino acid site in ß-glucuronidase. All our findings not only developed some potent novel ß-glucuronidase inhibitors but also indicated the potential herb drug interaction (HDI) effects of flavonoids with some clinical drugs which had enterohepatic circulation and further revealed the vital pharamcophoric requirement of natural flavonoids for ß-glucuronidase inhibition activity.

Two pairs of phenolic enantiomers from the leaves of Eucommia ulmoides Oliver.

Two pairs of new phenolic enantiomers, (+)-eucophenolic A (1a), (-)-eucophenolic B (1b), (-)-eucophenolic C (2a), (+)-eucophenolic D (2b) were isolated from the leaves of Eucommia ulmodies Oliver by chiral enantiomeric resolution. Their structures were elucidated based on extensive spectroscopic analysis. The absolute configurations of 1a/1b and 2a/2b were determined by empirical method and the calculated ECD and OR. All compounds were tested for Hep G2 tumour cell lines. However, no compounds showed potential cytotoxic activities against Hep G2 in vitro.

9-O-benzoyl-substituted berberine exerts a triglyceride-lowering effect through AMPK signaling pathway in human hepatoma HepG2 cells.

Berberine is an isoquinoline alkaloid extracted from Rhizoma coptidis and shows anti-hyperlipidemia effect in vivo and in vitro. We previously found that berberine could decrease the intracellular triglyceride content in human hepatoma HepG2 cells through activation of AMP-activated protein kinase (AMPK), a major regulator of lipid metabolism. Herein, to find a more effective agent, several berberine analogues (A1-A13) were isolated and synthesized, and the triglyceride-lowering effects and potential mechanisms were investigated in HepG2 cells. Among these berberine analogues, 9-O-benzoyl-substituted berberine (A13) showed strong affinity to AMPK and significantly up-regulated the levels of phospho-Thr172 AMPK α subunit. Meanwhile, A13 reduced the cellular triglyceride levels. Furthermore, A13 could mediate the mRNA levels of downstream proteins involved in triglyceride synthesis and fatty acid oxidation of AMPK signaling pathway. These results suggested that A13 exerts a triglyceride-lowering effect via stimulation of AMPK pathway, which may be beneficial to regulate hyperlipidemia.

Nine pairs of megastigmane enantiomers from the leaves of Eucommia ulmoides Oliver.

Nine pairs of megastigmane enantiomers (1a/1b-9a/9b), comprising two new compounds (6S,9R)-blumenol C (7b), (6S,9S)-blumenol C (8b), two pairs of enantiomers (+)-(6R)-eucomegastigmane A (1a), (-)-(6S)-eucomegastigmane A (1b), (+)-(3S,4S)-eucomegastigmane B (5a), (-)-(3R,4R)-eucomegastigmane B (5b) isolated by chiral resolution firstly, and twelve known compounds, were isolated from the leaves of Eucommia ulmoides Oliver. Their structures were elucidated based on extensive spectroscopic analysis. Absolute configurations of the megastigmane enantiomers were assigned by comparing experimental ECD and OR with calculated ECD and OR. Docking-based virtual screening of all compounds showed that megastigmane enantiomers have weak intermolecular interactions with the binding site residues of angiotensin-converting enzyme (ACE) and angiotensin II type 1 receptor (AT1R).

Megastigmane glycosides from leaves of Eucommia ulmoides Oliver with ACE inhibitory activity.

Four new megastigmane glycosides, eucomegastigsides A-D (2, 3, 5 and 7), together with three known megastigmane glycosides, (6R, 7E, 9R)-9-hydroxy-4, 7-megastigmadien-3-one-9-O-[α-l-arabinopyranosyl-(l→6)-ß-d-glucopyranoside (1), foliasalacioside B1 (4) and eleganoside A (6), were isolated from the leaves of Eucommia ulmoides Oliver. Their anti-hypertensive effect was investigated in vitro based on the inhibition of Angiotensin Converting Enzyme (ACE) using HPLC. The results showed that the isolates (2, 3, 4, 5, 7) had moderate inhibitory effects on ACE in vitro compared with captopril.

Berberine metabolites could induce low density lipoprotein receptor up-regulation to exert lipid-lowering effects in human hepatoma cells.

Berberine (BBR) is an isoquinoline alkaloid isolated from several Chinese herbal medicines, such as Coptis chinensis, Berberis aristata, and Coptis japonica. It exhibits a lipid-lowering effect by up-regulating the hepatic low density lipoprotein receptor (LDLR) expression. However, the plasma concentration of BBR is very low after oral administration for the reason that BBR is poorly absorbed and rapidly metabolized. Therefore, it is hard to explain the pharmacological effects of BBR in vivo. Here, RT-PCR, Western blotting and Oil Red O staining were used to investigate the effects of four BBR metabolites on LDLR expression and lipid accumulation in human hepatoma Hep G2 cells. Our results suggested that BBR increased the LDLR mRNA and protein levels in a time- and dose-dependent manner. Four metabolites of BBR, jatrorrhizine, columbamine, berberrubine and demethyleneberberine, were found to be able to up-regulate LDLR mRNA and protein expression. Moreover, almost all the metabolites had potent effects on inhibiting cellular lipid accumulation. These results suggest that both BBR and its metabolites exhibit lipid-lowering effects by up-regulating LDLR expression, and BBR and its metabolites might be the in vivo active forms of BBR produced after oral administration. This study provides information to help us understand the mechanisms underlying the hypolipidemic effects of BBR in vivo.