Optimization of trypsin extraction technology of Allium cepa L. polysaccharide by response surface methodology and the antitumor effects through immunomodulation.

The trypsin-assisted extraction of polysaccharides from Allium cepa L. was optimized using the response surface methodology (RSM). The optimum extraction conditions were extraction temperature, extraction time, extraction pH, and enzyme amount of 37.16°C, 180 min, 8.57, and 5.16%, respectively. Under the optimized conditions, the yield of A. cepa L. polysaccharides (ACP) reached 9.69%, which was comparable with the predicted yield (9.73%). Mid- and high-dose ACP significantly inhibited the tumor growth (43.93%) and the tumor inhibition percentage (38.05%), which were more than 30%. The ACP could extend the survival time of H22 ascites tumor-bearing mice. Furthermore, the ACP could reduce the thymus and the spleen atrophy and significantly promoted the Con A-induced proliferation of splenocytes and elevated the serum IFN-γ and IL-2 levels. Therefore, the ACP could inhibit the tumor growth in tumor-bearing mice and regulated the immune function of mice. Practical ApplicationsThe trypsin-assisted extraction has high efficiency, is carried out through the polysaccharide extraction and the deproteinization at the same time, and is more convenient and fast than traditional methods. No detailed study on the optimization of the trypsin extraction of onion polysaccharides is available. Thus, this experiment aims to use the BBD (4 factors and 3 levels) to optimize the roles of extraction temperature, extraction time, extraction pH, and amount of enzyme on the yield of polysaccharides obtained from the fruit of A. cepa L. In addition, when looking for high-quality biological functional principles for the pharmaceutical industry, the antitumor activity of ACP was evaluated. A. cepa L. is one of the most widely cultivated and consumed crops worldwide. Polysaccharides are the main active ingredient, and studies have shown that a high intake of Allium vegetables is associated with reduced risk of cancers.

A new benzophenone with biological activities from metabolites of butyrolactone I in rat faeces.

Butyrolactone I, one of the major secondary metabolites of fungus Aspergillus terreus, is a selective cdc2 kinase inhibitor. In the present study, the metabolism of butyrolactone I in male Wistar rats was investigated by characterising metabolites excreted into faeces. Following an oral dose of 40 mg/kg butyrolactone I, two phase I metabolites were isolated from the faeces of rat. The new structure was identified on the spectroscopic data analysis. The new compound V1 and known compound V2 were tested for their cytotoxicity, antimicrobial and antioxidant activity. V1 and V2 showed significant free radical scavenging ability. V2 also showed strong inhibitory activity against Staphylococcus aureus.

A new benzophenone with biological activities purified from Aspergillus fumigatus SWZ01.

Strain SZW01 was isolated from sea sediment collected from Shenzhen in Guangdong province, China, and was later identified as Aspergillus fumigatus by16S rDNA sequence analysis. Various chromatographic processes led to the isolation and purification of three compounds from the fermentation culture of SZW01, including a new compound, 2,6'-dihydroxy-2,4'dimethoxy-8'-methyl-6-methoxy-acyl-ethyl-diphenylmethanone (1), and two known compounds: fumigaclavine C (2) and alternarosin A (3), as characterised by UV, IR, 1 D, 2 D-NMR and MS data. The antioxidant and α-glucosidase inhibitory activities of these compounds were evaluated. The result illustrated that compound 1 exhibited a moderate antioxidant activity and stronger α-glucosidase inhibitory activity than acarbose.

KLF2 regulates neutrophil migration by modulating CXCR1 and CXCR2 in asthma.

Neutrophils are key inflammatory cells in the immunopathogenesis of asthma. Neutrophil migration can be initiated through activation of the CXCR1 and CXCR2 receptors by CXC chemokines, such as IL-8. Although transcription factor KLF2 has been found to maintain T cell migration patterns through repression of several chemokine receptors, whether KLF2 can regulate neutrophil migration via modulation of CXCR1 and CXCR2 is unknown. Here, we aimed to explore the functions of KLF2, CXCR1 and CXCR2 in neutrophil migration in asthma and to establish a regulatory role of KLF2 for CXCR1/2. We demonstrate that with asthma aggravation, the percentages and migration rates of peripheral blood neutrophils gradually increased in asthmatic patients and the guinea pig asthma model. Correspondingly, both the KLF2 mRNA and protein levels in neutrophils were gradually reduced. While CXCR1 and CXCR2 expression was negatively correlated with KLF2. In vitro knockdown of KLF2 dramatically increased the migration of HL-60-drived neutrophil-like cells, which was accompanied by an increase in the CXCR1 and CXCR2 mRNA and protein expression levels. Taken together, our results indicate that decreased KLF2 aggravates asthma progression by promoting neutrophil migration, which is associated with the transcriptional upregulation of CXCR1 and CXCR2. The KLF2 and/or CXCR1/2 expression levels may represent an indicator of asthma severity.

Sulfated modification, characterization and monosaccharide composition analysis of Undaria pinnatifida polysaccharides and anti-tumor activity.

Undaria pinnatifida (U. pinnatifida) polysaccharides (UPPS) are considered to be the major bioactive components of U. pinnatifida. The aim of the present study was to investigate the separation, sulfated modification, characterization and monosaccharide composition of UPPS. The optimal processing conditions were as follows: Distilled water-to-solid ratio, 50 ml/g; extraction time, 300 min; and extraction temperature, 90˚C. The major polysaccharide fraction of U. pinnatifida (UPPS-B1) was purified via DEAE-52 and Sephadex G-200 column chromatography. The chlorosulfonic acid-pyridine method was applied for sulfation modification. UPPS-B1 and sulfated (S)-UPPS-B1 were characterized via chemical analysis, ultraviolet-visible and Fourier-transformed infrared spectroscopy, gas chromatography and high-performance liquid chromatography. The total sugar content of UPPS-B1 and S-UPPS-B1 was 79.78 and 77.28%, respectively. The sulfate radical content of UPPS-B1 and S-UPPS-B1 was 8.53 and 29.12%, whilst the content of uronic acid was 9.29 and 7.98%, respectively. The average molecular weight of UPPS-B1 and S-UPPS-B1 was determined to be 37 and 110 kD, respectively. UPPS-B1 was considered to be a heteropolysaccharide composed of xylose, mannose, glucose and galactose at a ratio of 7.9:8.7:12.0:9.8. In addition, S-UPPS-B1 was a heteropolysaccharide composed of xylose, mannose, glucose and galactose at a ratio of 1.0:9.7:6.4:1.6. The results of the tumor growth inhibition experiment demonstrated that UPPS-B1 exhibited anti-tumor activity in vivo, which was improved following sulfation to yield S-UPPS-B1.

Enhancing in vivo oral bioavailability of cajaninstilbene acid using UDP-glucuronosyl transferase inhibitory excipient containing self-microemulsion.

Cajaninstilbene acid (CSA) exerts wide pharmacological activities, such as anti-inflammation, hypoglycaemic activity, analgesic effect and cognition improvement. However, it underwent severe phase II metabolism mediated by UDP-glucuronosyltransferase (UGT) in the gastrointestinal (GI) tract after oral administration, affecting its oral bioavailability. In the present study, we utilize UGT inhibitory excipient containing self-microemulsion (SME) delivery system to reduce the production of glucuronide metabolites and increase its oral bioavailability. The present results showed that although similar properties in physiochemical, cytotoxicity, cellular uptake, absorption and transport across rat everted gut sacs between SME-1 (inhibitory excipient containing SME) and SME-2 (control SME, without inhibitory excipient), an improved absolute bioavailability of 57.3 % was conferred by SME-1, significantly higher than the value of 35.4 % by SME-2 and 34.0 % by free CSA. Noticeably, the significantly lower AUC value of CSA glucuronide was determined in rats treated with SME-1 than those either treated with SME-2 or free CSA. Thus, the ability of SME-1 to enhance oral bioavailability of CSA is mainly attributed to the inhibition of phase II metabolism in the GI tract.

Antibacterial Diphenyl Ether, Benzophenone and Xanthone Derivatives from Aspergillus flavipes.

The extract of the strain Aspergillus flavipes DL-11 exerted antibacterial activities against six Gram-positive bacteria. During the following bioassay-guided separation, ten diphenyl ethers (1-10), two benzophenones (11-12), together with two xanthones (13-14) were isolated. Among them, 4'-chloroasterric acid (1) was a new chlorinated diphenyl ether. Their structures were elucidated by extensive spectroscopic data analysis, including IR, HR-ESI-MS, NMR experiments, and by comparison with the literature data. All compounds showed moderate to strong antibacterial effects on different Gram-positive bacteria with MIC values that ranged from 3.13 to 50 µg/mL, but none of the compounds exhibited activity against Gram-negative bacteria Vibrio parahaemolyticus ATCC17802 (MIC>100 µg/mL). In particular, the MICs of some compounds are at the level of positive control.

[Protective effect of raspberry extract on ConA-induced acute liver injuryin mice].

In this experiment,the antioxidant capacity of raspberry extract and the protective effect on liver injury induced by ConA in mice were investigated. Balb/C male mice were randomly divided into six groups: normal group,model group,bicyclol control group( 200 mg·kg~(-1)),low-dose raspberry extract group( 200 mg·kg~(-1)),middle-dose raspberry extract group( 400 mg·kg~(-1)),and highdose raspberry extract group( 800 mg·kg~(-1)). Each group was intragastrically administered with drugs according to the body weight once a day. Seven days later,all of the groups except for the normal group were treated with ConA( 20 mg·kg~(-1)) through tail vein injection to establish the acute liver injury model. The mice were put to death 8 hours later. The organ indexes were calculated. These rum levels of ALT,AST and LDH and the activities of SOD,CAT,GSH and MDA in liver tissue were detected. HE staining was used to observe the pathological changes of liver tissue in mice. Western blot was used to detect the expressions of Bax,Bcl-2,Nrf2 and Keap-1. The antioxidant capacity of raspberry extract was measured by CAA assay. The results showed that,raspberry extract had a strong antioxidant capacity. Simultaneously,compared with the model group,raspberry extract can significantly improve the pathological conditions of liver,and significantly reduce ALT,AST and LDH activities in serum of liver injury mice( P<0. 01). The activities of SOD,CAT in liver homogenate supernatant were significantly increased in the high-dose group,the content of GSH increased,while the content of MDA was sharply declined in the high-dose group( P<0. 01). Meanwhile,raspberry extract down-regulated the expressions of Bax and Keap-1 and up-regulated the expressions of Bcl-2 and Nrf2. CAA showed that the compound raspberry extract had a strong antioxidant capacity. Therefore,raspberry extract has an obvious protective effect on acute liver injury induced by ConA in mice.

Characterization of estrogenic active ingredients in Cuscuta chinensis Lam. based on spectral characteristics and high­performance liquid chromatography/quadrupole time­of­flight mass spectrometry.

High-performance liquid chromatography (HPLC) is an efficient method that is widely used to assess the quality of traditional Chinese medicine (TCM). It is well known that the quality of TCM has a direct effect on its efficacy; therefore, in order to thoroughly explain how TCM exerts its efficacy, it is necessary to characterize its active ingredients and assess their quality. The application of the spectrum­effect method is crucial for determining the pharmacological basis of materials. The aim of the present study was to examine the correlation between chemical spectra and estrogenic activity of Cuscuta chinensis Lam., in order to reveal active compounds with potential therapeutic effects. The spectra of Cuscuta chinensis Lam. were recorded using HPLC, and estrogenic activity was determined using a uterus growth test and MTT assay. Combination of the results of bivariate analysis, principal component analysis and Gray relational analysis identified 19 active compounds, as follows: Quercetin­3­O­(2'­O­α­rhamnosy­6'­O­malony)­â€‹ß­D­glucoside, ka-empferol­3­O­ß­D­aplosyl­(1→2)­â€‹[­α­â€‹L­rhamnosy­(1→6)]­ß-wD-glucoside, 6­O­(E)­P­coumaroyl)­ß­â€‹D­fructofuranosyl­(2→1)­α­D­glucopyranoside, kaempferol­7­rhamnosy, kaempferol­3­ß­D-glucuronide, apigenin, 4­caffeoyl­5­coumaroylquinic acid, kaempferol­3­arabofuranoside, quercetin­3­O-ß­D-apiofuranosyl-(1→2)-ß­D­galactoside, dicaffeoylquinic acid, hyperin, quercitin, isorhamnetin, chlorogenic acid, quercetin, quercltrin­2''­gallate, quercetin­3, 7­α­L­dirhamnoside and stigmasterol, as well as one unknown compound. The present study laid a foundation for in vivo metabolic studies regarding Cuscuta chinensis Lam. and for the development of its clinical application.

[Trace phenolic compounds from Red Yeast Rice].

Trace chemical constituents from the ethyl acetate extract of Red Yeast Rice were investigated. Four phenolic compounds were isolated by various column chromatographies, and their structures were identified on the basis of spectroscopic analysis including UV, MS, IR and NMR. The four compounds were identified as 2-methyl-5-(2'R-methyl-4'-hydroxy-butyl)-cinnamic acid(1), 5-(2'-hydroxy-6'-methyl phenyl)-3-methylfuran-2-carboxylic acid(2), daidzein(3), and genistein(4). Compound 1 was new and 2 was firstly discovered from the genus Monascus, while 3-4 were obtained from Red Yeast Rice for the first time.

Cytisine induces apoptosis of HepG2 cells.

Cytisine is a quinolizidine alkaloid, which has been reported to be among the major bioactive components of Sophora alopecuraides L. Quinolizidine alkaloids have previously been demonstrated to inhibit the proliferation of several types of tumor cells. However, few studies have investigated the effects of cytisine on cancer cells. The present study was performed to further investigate the molecular mechanisms underlying cytisine­induced apoptosis of HepG2 human hepatocellular carcinoma cells. The results of an MTT assay demonstrated that cytisine inhibited the growth of HepG2 cells in a dose­dependent manner. In addition, the induction of apoptosis was detected, as determined by morphological observation and flow cytometry. As determined by fluorescence microscopy, apoptotic morphological alterations were detected following cytisine administration. Flow cytometric analyses demonstrated that cytisine induced cytotoxicity through apoptosis­like mechanisms in HepG2 cells. Furthermore, western blot analysis was performed to investigate the release of cytochrome c (Cyt­c) and activation of the caspase cascade, and the results indicated that treatment of HepG2 cells with cytisine induced caspase­dependent apoptosis via the release of Cyt­c from the mitochondria, upregulation of caspase­3 and downregulation of pro­caspase­3. These results indicated that cytisine may induce apoptosis of HepG2 cells through the mitochondrial pathway.

A pair of new isocoumarin epimers from soil fungus Hypoxylon sp.

In our research on novel secondary metabolites from micro-organisms, two new (1-2) and four known dihydroisocoumarins (3-6) were derived from soil fungus Hypoxylon sp. Their structures were determined with extensive NMR data analysis and ECD calculation comparing with those of experimental CD spectra. Interestingly, compounds 1 and 2 possessed the same planar structure and very similar NMR data, suggesting 1 and 2 were a pair of epimers at either C-3 or at C-4, confirmed by the totally opposite cotton effect around 250 nm in the CD spectra of 1 and 2. Moreover, for the first time, we revealed that the CD absorption peak at 250 nm was dominated by C-3 orientation, rather than the orientation of C-3 substituents, by intensive ECD investigations.

[Study on the Apoptotic Mechanisms of Human Liver Cancer HepG-2 Cells Induced by Total Saponins of Ornithogalum caudatum].

Objective: To investigate the mechanism of human liver cancer HepG-2 cells apoptosis induced by total saponins of Ornithogalum caudatum( OCA-TS). Methods: The anti-proliferative effects of OCA-TS on HepG-2 cells was detected by MTT assay; the inverted microscope was used to observe cell morphology; transmission electron microscope( TEM) was used to observe the cellular ultrastructure changes; flow cytometry method was used to detect the apoptosis rate, mitochondrial membrane potential, and protein expression level of Cyt-C; Caspase-3 activity was measured by ELISA. Results: OCA-TS can inhibit the proliferation of HepG-2 cells and the IC50 was 79. 80 ± 0. 18 µg / m L. After treated by OCA-TS, cells became round, and the refractivity of cells receded, the number of suspension cells increased. By TEM method, the cells presented typical apoptosis characteristics. With the increasing of concentration of OCA-TS, cell apoptosis rate, the protein expression level of Cyt-C and the activity of Caspase-3 were increased markedly( P < 0. 05 or P < 0. 01). Conclusion: OCA-TS can effectively inhibit the proliferation of human liver cancer HepG-2 cells by inducing apoptosis of HepG-2 cells through mitochondrial pathway.

New flavonoid from Patrinia villosa.

CONTEXT: Patrinia villosa (Thunb.) Juss (Valerianaceae) is an important ancient herbal medicine widely used for inflammation, wound healing, and abdominal pain. But little is known of the phytochemical constituents of this herbal plant. OBJECTIVE: The objective of this study is to isolate and identify the bioactive components from P. villosa. MATERIALS AND METHODS: A 70% EtOH extract of P. villosa was subjected to normal-phase silica, ODS silica gel column chromatography, and semi-preparative HPLC chromatography after partitioned successively with light petroleum, dichloromethane and n-BuOH. Chemical structures of the compounds were elucidated by spectroscopic methods including UV, 1D-NMR, 2D-NMR, HR-ESI-MS, and CD spectra. The cytotoxic activity of the new component was determined with the SMMC-7721 cell line using the MTT method after incubation for 48 h. RESULTS: A new flavonoid named patriniaflavanone A (1) along with four known compounds was isolated from P. villosa. The four known compounds were identified as luteolin 7-O-glucuronide-6″-methyl ester (2), p-hydroxyphenylacetic acid methyl ester (3), trans-caffeic acid (4), and trans-caffeic acid methylate (5) by comparison of their spectral data with the reported data. The IC50 value of patriniaflavanone A (1) on SMMC-7721 was 61.27 µM. DISCUSSION AND CONCLUSION: This is the first report on the isolation and identification of patriniaflavanone A (1), and compounds 2-5 were isolated for the first time from the title plant. Patriniaflavanone A (1) exhibited moderate cytotoxic activity.

In vitro analysis of the role of the mitochondrial apoptosis pathway in CSBE therapy against human gastric cancer.

The caper plant (Capparis spinosa L.) was a common Uyghur folk medicine, and is a member of the Capparidaceae family. In a previous study, the n-butanol extract of C. spinosa L. (CSBE) was demonstrated to exert anti-tumor activity; however, the underlying mechanism is currently not understood. The present study aimed to elucidate the mechanism underlying the CSBE-induced mitochondrial apoptotic pathway, in order to investigate the anti-tumor effects of this plant extract. CSBE-induced apoptosis of the SGC-7901 human gastric cancer cell line was observed, and alterations in the expression levels and localization of initiators, markers, and executors of the mitochondrial apoptosis pathway were analyzed. Following treatment of SGC-7901 cells with CBSE, proliferation was inhibited and apoptosis was induced; and these effects were associated with mitochondrial membrane potential disruption, cytochrome c release into the cytoplasm, and caspase-9 and caspase-3 activation. CSBE may have induced SGC-7901 cell apoptosis by upregulating the expression of B-cell lymphoma-2 (BCL-2)-associated X protein, and downregulating the expression of BCL-2. The results of the present study suggested that CSBE may induce SGC-7901 cell apoptosis via activation of the mitochondrial apoptosis pathway.

Magnesium ions improving the growth and organics reduction of Rhodospirillum rubrum cultivated in sewage through regulating energy metabolism pathways.

Rhodospirillum rubrum has the potential for biomass resource recycling combined with sewage purification. However, low biomass production and yield restricts the potential for sewage purification. This research investigated the improvement of biomass production, yield and organics reduction by Mg²âº in R. rubrum wastewater treatment. Results showed that with optimal dosage (120 mg/L), biomass production reached 4,000 mg/L, which was 1.5 times of that of the control group. Biomass yield was improved by 43.3%. Chemical oxygen demand (COD) removal reached over 90%. Hydraulic retention time was shortened by 25%. Mechanism analysis indicated that Mg²âº enhanced the isocitrate dehydrogenase and Ca²âº/Mg²âº-ATPase activities, bacteriochlorophyll content on respiration and photophosphorylation. These effects then enhanced ATP production, which led to more biomass accumulation and COD removal. With 120 mg/L Mg²âº dosage, the isocitrate dehydrogenase and Ca²âº/Mg²âº-ATPase activities, bacteriochlorophyll content, ATP production were improved, respectively, by 33.3%, 50%, 67%, 41.3% compared to those of the control group.

New C13 lipids from the marine-derived fungus Trichoderma harzianum.

Chemical examination of the fermentation broth of a sponge-associated fungus Trichoderma harzinum HMS-15-3 led to the isolation of four pairs of new C13 lipid enantiomers namely harzianumols A-H (1a-4b). Their structures were elucidated on the basis of extensive spectroscopic (IR, MS, 1D, and 2D NMR) data analysis, including the modified Mosher's method for the assignment of their absolute configurations. The new compounds were evaluated for antihyperlipidemic effects in HepG2 cells.

Targeted gene capture and massively parallel sequencing identify TMC1 as the causative gene in a six-generation Chinese family with autosomal dominant hearing loss.

Hereditary nonsyndromic hearing loss is extremely heterogeneous. Mutations in the transmembrane channel-like gene1 (TMC1) are known to cause autosomal dominant and recessive forms of nonsyndromic hearing loss linked to the loci of DFNA36 and DFNB7/11, respectively. We characterized a six-generation Chinese family (5315) with progressive, postlingual autosomal dominant nonsyndromic hearing loss (ADNSHL). By combining targeted capture of 82 known deafness genes, next-generation sequencing and bioinformatic analysis, we identified TMC1 c.1714G>A (p. D572N) as the disease-causing mutation. This mutation co-segregated with hearing loss in other family members and was not detected in 308 normal controls. In order to determine the prevalence of TMC1 c.1714G>A in Chinese ADNSHL families, we used DNA samples from 67 ADNSHL families with sloping audiogram and identified two families carry this mutation. To determine whether it arose from a common ancestor, we analyzed nine STR markers. Our results indicated that TMC1 c.1714G>A (p.D572N) account for about 4.4% (3/68) of ADNSHL in the Chinese population.

Screening antitumor bioactive fraction from Sauromatum giganteum (Engl.) Cusimano & Hett and sensitive cell lines with the serum pharmacology method and identification by UPLC-TOF-MS.

Sauromatum giganteum (Engl.) Cusimano & Hett Tuber are used in Chinese folklore medicine for treatment of neoplasms. However, the claim has not been scientifically validated. The aim of the study is to screen the antitumor bioactive fraction of Sauromatum giganteum (Engl.) Cusimano & Hett Tuber and sensitive tumor cell lines using a cytotoxicity assay in vitro and tumor transplantation method in vivo, to support its use in folk medicine. The petroleum ether fraction, chloroform fraction, ethyl acetate fraction, n-butanol fraction and water fraction were successively extracted by turn by the maceration under reflux assay. Screening of antitumor bioactive fraction and sensitive cell lines were measured by MTT assay and the serum pharmacology method, and in vivo the antitumor activities of the active fraction was evaluated by using S180 or H22 tumor-bearing mice model and Kunming mice. The active constituents of ethyl acetate fraction of Sauromatum giganteum (Engl.) Cusimano & Hett were characterized by UPLC-TOF-MS. Compared with control groups, mice serum containing ethyl acetate fraction had a inhibition effect on SMMC-7721 cell, SGC-7901 cell, MCF-7 cell, HeLa cell, A549 cell, HT-29, and MDA-MB-231, respectively, but mice serum containing other four fractions had no different with that of control group. The inhibition capabilities of mice serum containing ethyl acetate fraction on the seven cell lines in descending order is SGC-7901 > SMMC-7721 > MCF-7 > HT-29 > A549 > HeLa > MDA-MB-231. In vivo the inhibition rate of 106, 318, 954 mg/kg·d ethyl acetate fraction dry extract to sarcoma S180 is 15.22%, 26.15% and 40.24%, respectively, and life prolonging rate to hepatoma H22 is 33.61%, 40.16% and 55.74%. A total of 14 compounds were identified in the ethyl acetate fraction of Sauromatum giganteum (Engl.) Cusimano & Hett. The results of the experimental studies proved the antitumor activity of Sauromatum giganteum (Engl.) Cusimano & Hett and supported the traditional use of this plant. These data indicate the potential for the use of ethyl acetate fraction of Sauromatum giganteum (Engl.) Cusimano & Hett Tuber in tumor therapy, anti-tumor activity on cancer cell line in descending order is SGC-7901 > SMMC-7721 > MCF-7 > HT-29 > A549 > HeLa > MDA-MB-231.

Active components alignment of Gegenqinlian decoction protects ulcerative colitis by attenuating inflammatory and oxidative stress.

ETHNOPHARMACOLOGICAL RELEVANCE: Gegenqinlian Decoction (GQD) has been used as a folk remedy for gastrointestinal diseases in China over thousands of years. It has significant treatment efficacy for patients with inflammatory bowel disease (IBD). We analyzed and showed that the active components alignment of Gegenqinlian Decoction (ACAG) possesses broad pharmacological effects including analgesic, antipyretic, anti-inflammatory, antibacterial, antiviral and antidiarrhea, as well as the effect of adjusting gastrointestinal function in our preliminary experiments. However, the exact molecular mechanisms on how ACAG exerts these pharmacological effects still remain elusive. In the present study, the plausible pharmacological effects of ACAG on 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis were investigated. MATERIALS AND METHODS: Male Sprague-Dawley (SD) rats with TNBS/ethanol-induced colitis were used. The colonic wet weight, macroscopic and histological colon injury, superoxide dismutase (SOD), malonyldialdehyde (MDA), and inducible nitric oxide synthase (iNOS) activity were observed. Pro-inflammation cytokines were determined by ELISA methods, semi-quantitative RT-PCR and Immuno-histochemistry. RESULTS: We showed administration of ACAG was able to improve colitis. This was manifested by a decreased in the score of macroscopic and histological colonic injury, by lowered colonic wet weight, accompanied by significant increased of SOD activity, and decreased of MDA and iNOS activities. The treatment also significantly reduced tumor necrosis factor-alpha (TNF-α) and interleukin-1ß (IL-1ß) levels in colon and serum as well as the colonic mRNA levels for several inflammatory cytokines such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), macrophage inflammatory protein-2 (MIP-2), intercellular adhesion molecule-1 (ICAM-1) and toll-like receptor 2, 4 (TLR2, TLR4). In addition, we also showed that ACAG was able to inhibit the activation and translocation of transcription factors, nuclear factor kappaBp65 (NF-κBp65) in colon. CONCLUSIONS: Our results suggest that ACAG exhibits protective effect in TNBS-induced ulcerative colitis. We postulate that this might be due to its modulation of oxidant/anti-oxidant balance, downregulation of productions, expressions of pro-inflammatory cytokines and inhibition of NF-κBp65 signal transduction pathways.