Synthesis and In Vitro Evaluation of Caffeoylquinic Acid Derivatives as Potential Hypolipidemic Agents.

A series of novel caffeoylquinic acid derivatives of chlorogenic acid have been designed and synthesized. Biological evaluation indicated that several synthesized derivatives exhibited moderate to good lipid-lowering effects on oleic acid-elicited lipid accumulation in HepG2 liver cells. Particularly, derivatives 3d, 3g, 4c and 4d exhibited more potential lipid-lowering effect than the positive control simvastatin and chlorogenic acid. Further studies on the mechanism of 3d, 3g, 4c and 4d revealed that the lipid-lowering effects were related to their regulation of TG levels and merit further investigation.

[Pandanus tectorius derived caffeoylquinic acids inhibit lipid accumulation in HepG2 hepatoma cells through regulation of gene expression involved in lipid metabolism].

The fruit of Pandanus tectorius (PTF) has a long history of use as a folk medicine to treat hyperlipidemia in Hainan province, South China. Our previous studies have shown that the n-butanol extract of PTF is rich in caffeoylquinic acids and has an adequate therapeutic effect on dyslipidemic animals induced by high-fat diet. In this work, seven caffeoylquinic acids isolated from PTF were screened for the lipid-lowering activity in HepG2 hepatoma cells. Oil-Red O staining, microscopy and intracellular triglyceride (TG) and total cholesterol (TC) quantification showed that 3-O-caffeoylquinic acid (3-CQA), 3, 5-di-O-caffeoylquinic acid (3,5-CQA), and 3,4,5-tri-O-caffeoylquinic acid (3,4,5-CQA) significantly inhibited lipid accumulation induced by oleic acid and decreased intracellular levels of TC and TG in a dose-dependent manner. These three caffeoylquinic acids showed no significant cytotoxicity at concentrations of 1 -50 µmol x L(-1) as determined by MTT assay. Realtime quantitative PCR revealed that 3-CQA and 3, 5-CQA significantly increased the expression of lipid oxidation-related genes PPARα, CPT-1 and ACOX1 while 3-CQA, 3, 5-CQA and 3,4,5-CQA decreased the expression of lipogenic genes SREBP-1c, SREBP-2, HMGR, ACC, FAS. Overall, 3-CQA, 3, 5-CQA and 3, 4, 5-CQA may be the principal hypolipidemic components in PTF which can decrease intracellular lipid accumulation through up-regulating the expression of lipid oxidative genes and down-regulating the expression of lipogenic genes.

From ancient wisdom to modern science: Gut microbiota sheds light on property theory of traditional Chinese medicine.

The property theory of traditional Chinese medicine (TCM) has been practiced for thousands of years, playing a pivotal role in the clinical application of TCM. While advancements in energy metabolism, chemical composition analysis, machine learning, ion current modeling, and supercritical fluid technology have provided valuable insight into how aspects of TCM property theory may be measured, these studies only capture specific aspects of TCM property theory in isolation, overlooking the holistic perspective inherent in TCM. To systematically investigate the modern interpretation of the TCM property theory from multidimensional perspectives, we consulted the Chinese Pharmacopoeia (2020 edition) to compile a list of Chinese materia medica (CMM). Then, using the Latin names of each CMM and gut microbiota as keywords, we searched the PubMed database for relevant research on gut microbiota and CMM. The regulatory patterns of different herbs on gut microbiota were then summarized from the perspectives of the four natures, the five flavors and the meridian tropism. In terms of the four natures, we found that warm-natured medicines promoted the colonization of specific beneficial bacteria, while cold-natured medicines boosted populations of some beneficial bacteria while suppressing pathogenic bacteria. Analysis of the five flavors revealed that sweet-flavored and bitter-flavored CMMs positively influenced beneficial bacteria while inhibiting harmful bacteria. CMMs with different meridian tropism exhibited complex modulative patterns on gut microbiota, with Jueyin (Liver) and Taiyin (Lung) meridian CMMs generally exerting a stronger effect. The gut microbiota may be a biological indicator for characterizing the TCM property theory, which not only enhances our understanding of classic TCM theory but also contributes to its scientific advancement and application in healthcare. Please cite this article as: Yang YN, Zhan JG, Cao Y, Wu CM. From ancient wisdom to modern science: Gut microbiota sheds light on property theory of traditional Chinese medicine. J Integr Med 2024; 22(4): 413-445.

Oral bacteriome and mycobiome of patients with idiopathic membranous nephropathy with different tongue coatings treated with a Chinese herbal formula.

ETHNOPHARMACOLOGICAL RELEVANCE: Moshen Fuyuan Formula (MSFY) is one of the representative Chinese medicine compound for Idiopathic membranous nephropathy (IMN), that originate from Fang Ji Huang Qi decoction in the Han dynasty. IMN is usually accompanied by different tongue coatings in traditional Chinese medicine (TCM), and tongue microorganisms are important factors affecting the formation of the tongue coating. Recently, oral microbiomes, including bacteria and fungi, have been identified as pivotal factors that contribute to disease development. However, the regulation of oral microbiomes by MSFY has not been defined. AIM OF THE STUDY: In this work, we explore the characteristics of oral bacteria and fungi in IMN patients with different tongue coatings, and clarify the therapeutic effect of MSFY based on oral microbiome. MATERIALS AND METHODS: We enrolled 24 patients with IMN, including 11 with white tongue (IMN-W) and 13 with yellow tongue (IMN-Y), and recruited an additional 10 healthy individuals. Patients with IMN were treated with the MSFY. The oral bacteriome and fungi before and after treatment were detected using full-length 16S rRNA and internal transcribed spacer gene sequencing. RESULTS: The therapeutic effect of MSFY on patients with yellow tongue coating was more significant than that on patients with white tongue coating. In terms of oral bacteriome, Campylobacter bacteria were enriched in patients with yellow tongue and could be a promising biomarker for yellow coating. Enrichment of Veillonella parvula_A may partially account for the therapeutic effect of MSFY. As for oral fungi, Malassezia globosa was enhanced in patients with IMN-W and reduced in patients with IMN-Y. Notably, it was reduced by MSFY. We also found that mycobiome-bacteriome interactions were highly complex and dynamic in patients with IMN. CONCLUSION: The regulation of the dynamic balance between oral fungi and bacteria by MSFY contributes to the treatment of IMN. This study determined the oral bacteriome and mycobiome of patients with IMN with different tongue coatings before and after MSFY treatment, which aids in promoting personalized treatment in clinical TCM and provides direction for investigating the mechanism of Chinese herbal medicines.

The gut microbiota-aromatic hydrocarbon receptor (AhR) axis mediates the anticolitic effect of polyphenol-rich extracts from Sanghuangporus.

Inflammatory bowel disease (IBD) is a significant global health concern. The gut microbiota plays an essential role in the onset and development of IBD. Sanghuangporus (SH), a traditional Chinese medicinal mushroom, has excellent anti-inflammatory effects and is effective at modulating the gut microbiota. Despite these attributes, the specific anticolitic effects of SH and the mechanisms through which the gut microbiota mediates its benefits remain unclear. Herein, we demonstrated that polyphenol-rich extract from SH effectively alleviated the pathological symptoms of dextran sodium sulfate (DSS)-induced colitis in mice by modulating the gut microbiota. Treatment with SH distinctly enriched Alistipes, especially Alistipes onderdonkii, and its metabolite 5-hydroxyindole-3-acetic acid (5HIAA). Oral gavage of live A. onderdonkii or 5HIAA potently mitigated DSS-induced colitis in mice. Moreover, both 5HIAA and SH significantly activated the aromatic hydrocarbon receptor (AhR), and the administration of an AhR antagonist abrogated their protective effects against colitis. These results underscore the potent efficacy of SH in diminishing DSS-induced colitis through the promotion of A. onderdonkii and 5HIAA, ultimately activating AhR signaling. This study unveils potential avenues for developing therapeutic strategies for colitis based on the interplay between SH and the gut microbiota.

Development of a rapid multi-residue assay for detecting ß-lactams using penicillin binding protein 2x*.

OBJECTIVE: To develop a rapid multi-residue assay for detecting 16 demanded by the European Union (EU). METHODS: A recombinant penicillin-binding protein (PBP) 2x* from Streptococcus pneumoniae R6 was expressed in vitro and six ß-lactams were conjugated to HRP by four methods. A rapid multi-residue assay for ß-lactams was established with PBP2x* and HRP-conjugate. RESULTS: PBP2x* was expressed and purified successfully and the ideal HRP-conjugate was identified. The multi-residue assay was developed. After optimization, penicillin G, ampicillin, amoxicillin, cloxacillin, dicloxacillin, oxacillin, nafcillin, cephalexin, ceftiofur, cefalonium, cefquinome, cefazolin, cefoperazone, cephacetrile, and cephapirin can be detected at levels below MRL in milk with simple pretreatment. CONCLUSION: This assay developed can detect all 16 ß-lactams demanded by the European Union (EU). The whole procedure takes only 45 min and can detect 42 samples and the standards with duplicate analysis.

Structural modulation of gut microbiota during alleviation of experimental passive Heymann nephritis in rats by a traditional Chinese herbal formula.

BACKGROUND: Jianpi-Qushi-Heluo formula (JQHF) has been used to treat idiopathic membranous nephropathy (IMN) in hospitals for many years. PURPOSE: Elucidating the protective effect and exploring the potential mechanism of JQHF against IMN. METHODS: Passive Heymann nephritis (PHN) was induced in rats by a single tail vein injection of anti-Fx1A antiserum. Then, the animals were treated with JQHF at 16.2 g/kg or 32.4 g/kg, with benzepril (10 mg/kg) as a positive control. Renal function was evaluated by biochemical measurements and pathological testing. Fecal samples were collected before and after treatment to analyze the gut microbiota composition by shotgun whole metagenome sequencing. RESULTS: JQHF exhibited potent efficacy in ameliorating PHN at both doses, as revealed by decreasing the deposition of IgG and C5b-9, relieving podocyte injury, and reducing glomerular and tubular cell apoptosis. The lower dose was corresponding to the clinical dosage and showed better therapeutic effects than the higher dose. Metagenomic analysis showed that gavage with 16.2 g/kg of JQHF shifted the structure of the gut microbiota in PHN rats and significantly increased the relative abundances of Prevotella copri, Lactobacillus vaginalis and Subdoligranulum variabile. Particularly, S. variabile was strongly negatively correlated with serum levels of TC and TG, the deposition of IgG and C5b-9, and apoptosis of glomerular cells. CONCLUSIONS: The JQHF is an effective agent for the treatment of experimental PHN. The PHN-allevating effect of JQHF is associated with specific alternation of gut microbiota.

The Gut Microbial Co-Abundance Gene Groups (CAGs) Differentially Respond to the Flavor (Yao-Wei) of Chinese Materia Medica.

The property theory is a unique principle instructing traditional Chinese doctors to prescribe proper medicines against diseases. As an essential part of it, the five-flavor theory catalogs various Chinese materia medicas (CMMs) into five flavors (sweet, bitter, sour, salty, and pungent) based on their taste and medical functions. Although CMM has been successfully applied in China for thousands of years, it is still a big challenge to interpret CMM flavor via modern biomarkers, further deepening its elusiveness. Herein, to identify the correlation between gut microbiota and CMM flavor, we selected 14 CMMs with different flavors to prepare their aqueous extracts, quantified the contained major chemical components, and then performed full-length 16S rRNA sequencing to analyze the gut microbiota of C57BL/6 mice administrated with CMM extracts. We found that flavones, alkaloids, and saponins were the richest components for sweet-, bitter-, and pungent-flavored CMMs, respectively. Medicines with merged flavors (bitter-pungent and sweet-pungent) displayed mixed profiles of components. According to gut microbial analysis, modulation of CMMs belonging to the same flavor on the taxonomic classification was inconsistent to an extent, while the functional sets of gut microbiota, co-abundance gene groups (CAGs), strongly and differentially responded to distinct flavors. Moreover, these correlations were in line with their pharmacological actions. Therefore, the gut microbial functional sets (CAGs) could act as the possible indicator to reflect CMM flavor, rather than the composition of microbial community.

Cordycepin prevents hyperlipidemia in hamsters fed a high-fat diet via activation of AMP-activated protein kinase.

Hyperlipidemia is a major risk factor for cardiovascular diseases. In this study, we investigated the potential effects of cordycepin (3'-deoxyadenosine), a bioactive component of the fungus Cordyceps militaris, on hyperlipidemia. We found that in male Syrian golden hamsters fed a high-fat diet (HFD), daily administration of cordycepin effectively reduced the accumulation of serum total cholesterol (TC), triglycerides (TG), and low-density lipoprotein cholesterol (LDL-c) and suppressed HFD-associated increases in relative retroperitoneal fat. It also increased the levels of phospho-AMP-activated protein kinase (AMPK) and phospho-acetyl-CoA carboxylase (phospho-ACC) in liver and retroperitoneal adipose tissues. In HepG2 cells, cordycepin stimulated robust concentration- and time-dependent AMPK activation that correlated with the activation of ACC and the suppression of lipid biosynthesis. However, pretreatment with compound C, a specific inhibitor of AMPK, substantially abolished the effects of cordycepin on AMPK activation and lipid biosynthesis inhibition. These results indicate that cordycepin prevents hyperlipidemia via activation of AMPK. Experiments on abnormal metabolic mice indicated that cordycepin can also improve insulin sensitivity effectively.

The adenosine derivative 2',3',5'-tri-O-acetyl-N6-(3-hydroxylaniline) adenosine activates AMPK and regulates lipid metabolism in vitro and in vivo.

AIMS: Our overall objective was to investigate the effect of the adenosine derivative 2',3',5'-tri-O-acetyl-N6-(3-hydroxylaniline) adenosine (WS010117) on AMP-activated protein kinase (AMPK) activation and lipid metabolism and to also assess the underlying mechanisms involved in these processes. MAIN METHODS: HepG2 cells and hamsters fed a high-fat diet were used to test the effects of WS010117 on lipid metabolism. Western blots, chemical intervention, HPLC, SAMS peptide assay, (14)C-labelled acetate and palmitate assays, molecular docking assay and siRNA targeting the AMPK γ1 subunit were used to investigate the effect of WS010117 on AMPK activation as well as the underlying mechanism involved in this activation. KEY FINDINGS: WS010117 treatment resulted in the dose-dependent activation of AMPK in HepG2 cells, increasing lipid oxidation and decreasing lipid biosynthesis. In hamsters that were fed a high-fat diet, WS010117 treatment (1.5-6 mg/kg) significantly inhibited the increase in lipid accumulation. WS010117-induced AMPK activation was essentially abolished by treatment with compound C, and the addition of WS010117 did not alter the intracellular AMP:ATP ratio. In HeLa cells endogenously lacking LKB1, WS010117-mediated AMPK activation was not impaired, even following co-treatment with STO-609, a selective inhibitor of Ca(2+)/calmodulin-dependent protein kinase kinase (CaMKK). The results from the molecular docking assays and experiments targeting the AMPK γ1 subunit with siRNA indicated that WS010117 may activate AMPK by binding to and regulating the γ subunit of AMPK. SIGNIFICANCE: Our data indicate that WS010117 can regulate lipid metabolism through the activation of AMPK. WS010117 may activate AMPK by binding to and regulating the AMPK γ subunit.

Combination of HPLC chromatogram and hypoglycemic effect identifies isoflavones as the principal active fraction of Belamcanda chinensis leaf extract in diabetes treatment.

In previous study, we demonstrated the hypoglycemic effect of aqueous extract of Belamcanda chinensis leaves in rats. Here, we separated the aqueous extract of B. chinensis leaves and investigated the spectrum-effect relationships between HPLC chromatograms and hypoglycemic activities of different isolates from B. chinensis leaf extract. Sequential solvent extraction with petroleum ether, chloroform, acetic ester and n-butanol provided several isolates showing similar hypoglycemic activities, making it difficult to discriminate the active fractions. Stepwise elution through HP20 macroporous resin by water, 40% and 95% ethanol provided isolates with distinct hypoglycemic activities, representing a simple, rapid and efficient preparative separation method. Combination of HPLC chromatogram and pharmacological effect targeted a hypoglycemic activity-related region in HPLC chromatogram. Each peak in this region was analyzed by UV spectrum scan. Most of them were flavonoids in which tectoridin and swertisin were known flavonoids with anti-diabetic activities. In together, this work provides a general model of combination of HPLC chromatography and pharmacological effect to study the spectrum-effect relationships of aqueous extract from B. chinensis leaves, which can be used to find principle components of B. chinensis on pharmacological activity.

Antihyperglycemic effect of Cephalotaxus sinensis leaves and GLUT-4 translocation facilitating activity of its flavonoid constituents.

The objectives of this study were to investigate the antihyperglycemic effect of Cephalotaxus sinensis leaves and to identify the active components. The antihyperglycemic effect of various fractions (FA, FB, FC, FD) of the 80% ethanol extract of the leaves was evaluated in streptozotocin (STZ)-induced diabetic rats. Among the tested fractions, FC was the most active. FC (0.48 g/kg) given orally for 10 d reduced significantly (p<0.001) the blood glucose of STZ-induced diabetic rats. The food and water intakes of FC (0.48 g/kg)-treated diabetic rats were reduced significantly (p<0.001) when compared to the 0.5% carboxymethyl cellulose (CMC)-treated diabetic rats. The activity-guided fractionation of the ethanol extract of C. sinensis leaves furnished three flavonoid compounds, apigenin-5-O-[alpha-L-rhamnopyranosyl-(1-->4)-6-O-beta-D-acetylglucopyranoside] (1), apigenin (2), and apigenin-5-O-[alpha-L-rhamnopyranosyl-(1-->4)-6-O-beta-D-glucopyranoside] (3). The elevation of GLUT-4 protein level in membrane preparations from mice adipocytes was detected by Western blot analysis after adipocytes were pre-incubated with FC (0.1, 1, 10 mg/ml), apigenin (0.1, 2 mg/ml) and apigenin-5-O-[alpha-L-rhamnopyranosyl-(1-->4)-6-O-beta-D-acetylglucopyranoside] (0.1, 2 mg/ml), respectively. Phytochemical investigation and HPLC-DAD analysis of FC indicated that the flavonoids were the major constituents in this fraction. These results suggest that the fraction from C. sinensis leaves is a promising drug for the treatment of diabetes, and that the flavonoids from this plant are the active constituents.