Fishing of α-Glucosidase's Ligands from Aloe vera by α-Glucosidase Functionalized Magnetic Nanoparticles.

α-Glucosidase was immobilized on magnetic nanoparticles (MNPs) for selective solid-phase extraction of the enzyme's ligands present in Aloe vera, which is a medicinal plant used for the treatment of various diseases and possesses anti-diabetic activity. One new compound, aloeacone (2), together with two known compounds, aloenin aglycone (1) and aloin A (3), were fished out as the enzyme's ligands. The structure of 2 was determined by HR-MS and comprehensive NMR techniques. Compound 3 exhibited a weak inhibitory effect on α-glucosidase, while compounds 1 and 2 were found to possess activation effects on the enzyme for the first time. It is interesting that both an inhibitor and agonists of α-glucosidase were fished out in one experiment.

Cytosolic Acetyl-CoA Generated by ATP-Citrate Lyase Is Essential for Acetylation of Cell Wall Polysaccharides.

Plant cell wall polysaccharides, including xylan, glucomannan, xyloglucan and pectin, are often acetylated. Although a number of acetyltransferases responsible for the acetylation of some of these polysaccharides have been biochemically characterized, little is known about the source of acetyl donors and how acetyl donors are translocated into the Golgi, where these polysaccharides are synthesized. In this report, we investigated roles of ATP-citrate lyase (ACL) that generates cytosolic acetyl-CoA in cell wall polysaccharide acetylation and effects of simultaneous mutations of four Reduced Wall Acetylation (RWA) genes on acetyl-CoA transport into the Golgi in Arabidopsis thaliana. Expression analyses of genes involved in the generation of acetyl-CoA in different subcellular compartments showed that the expression of several ACL genes responsible for cytosolic acetyl-CoA synthesis was elevated in interfascicular fiber cells and induced by secondary wall-associated transcriptional activators. Simultaneous downregulation of the expression of ACL genes was demonstrated to result in a substantial decrease in the degree of xylan acetylation and a severe alteration in secondary wall structure in xylem vessels. In addition, the degree of acetylation of other cell wall polysaccharides, including glucomannan, xyloglucan and pectin, was also reduced. Moreover, Golgi-enriched membrane vesicles isolated from the rwa1/2/3/4 quadruple mutant were found to exhibit a drastic reduction in acetyl-CoA transport activity compared with the wild type. These findings indicate that cytosolic acetyl-CoA generated by ACL is essential for cell wall polysaccharide acetylation and RWAs are required for its transport from the cytosol into the Golgi.

Colonic transit: what is the impact of a diverting loop ileostomy?

BACKGROUND: Diverting loop ileostomy (DLI) is used following low anterior resections (LAR) or ultra-low anterior resections (ULAR) to reduce anastomotic leak (AL). Preoperative mechanical bowel preparation (MBP) is traditionally used with DLI. However, clearance of the left colon can be achieved with a fleet enema without the physiological compromise of MBP. We aimed to assess colonic transit following DLI in this context. METHODS: A prospective, observational study was performed with patients with rectal cancer undergoing LAR or ULAR in a tertiary colorectal unit with preoperative fleet enema. Radiopaque markers were inserted into the caecum following rectal resection and formation of a DLI with placement confirmed by image intensifier and endoscopy. X-rays were performed at days 1, 3, 5 and 14 post-operation with data collected prospectively. RESULTS: Ten patients (mean age 57, nine males) were enrolled. Mean time to functioning stoma was 1.9 days (range 1-3). There was no movement in the majority of markers in all patients at Day 5 post-operation. In all seven patients with Day 14 X-rays, the majority of markers remained in the right colon. Two patients had delayed AL, with markers found within the pelvis in both of these patients. CONCLUSIONS: This is the first study to assess colonic transit following DLI using fleet enema only, with results suggesting colonic motility is abolished in this setting. The use of a fleet enema without MBP may be sufficient prior to rectal resection surgery when DLI is employed. AL may actually increase colonic transit. Further research is warranted.

Sennoside B inhibits PDGF receptor signaling and cell proliferation induced by PDGF-BB in human osteosarcoma cells.

AIMS: To address the possibility that sennoside B inhibition of cell proliferation is mediated via interference with platelet-derived growth factor (PDGF) signaling. MAIN METHODS: Human osteosarcoma MG63 cells were treated with PDGF in the presence or absence of sennoside B. Activation of the PDGF signaling pathway was monitored using western immunoblotting with specific antibodies against the PDGF receptor, phosphotyrosine and components of the downstream signaling cascade. Activation of cell metabolism and proliferation was assessed by chromogenic reduction of MTT. KEY FINDINGS: Sennoside B was found to inhibit PDGF-BB-induced phosphorylation of the PDGF receptor (PDGFR) in human MG63 osteosarcoma cells. Downstream signaling was also affected; pre-incubation of PDGF-BB with sennoside B inhibited the phosphorylation of pathway components including Ak strain transforming protein (AKT), signal transducer and activator of transcription 5 (STAT-5) and extracellular signal-regulated kinase 1/2 (ERK1/2). Further, we found that sennoside B can bind directly to the extracellular domains of both PDGF-BB and the PDGF-beta receptor (PDGFR-beta). The effect was specific for sennoside B; other similar compounds including aloe-emodin, rhein and the meso isomer (sennoside A) failed to inhibit PDGFR activation or downstream signaling. Sennoside B also inhibited PDGF-BB stimulation of MG63 cell proliferation. SIGNIFICANCE: These results indicate that sennoside B can inhibit PDGF-stimulated cell proliferation by binding to PDGF-BB and its receptor and by down-regulating the PDGFR-beta signaling pathway. Sennoside B is therefore of potential utility in the treatment of proliferative diseases in which PDGF signaling plays a central role.

Low-dose lactulose produces a tonic contraction in the human colon.

Lactulose (10-20 g day(-1)) is used to treat constipation. At this therapeutic dose, its effects on colonic motility remain unknown. Twenty-two healthy subjects swallowed a probe with an infusion catheter, six perfused catheters and a balloon connected to a barostat. Colonic phasic and tonic motor activity was recorded in fasting state. In group 1, four volunteers ingested 15 g lactulose and motility was recorded for 5 h after entry of lactulose into the caecum; in group 2, motility was recorded during (3 h) and 2 h after intracolonic infusion of isoosmotic and isovolumetric solutions containing sodium chloride alone (n = 9) or with 15 g lactulose (n = 9). In a last group of volunteers, isotopic colonic transit after ingestion of lactulose (10 g,n = 9) was assessed and compared with a control group (n = 17). Ingestion or intracolonic infusion of 15 g lactulose significantly decreased barostat bag volume (maximal decrease: 45 +/- 12% and 35 +/- 9% of basal value respectively). Phasic contractions remained unchanged. Tonic and phasic motility was unchanged by the isotonic and isovolumetric infusion of saline. Ingestion of lactulose significantly accelerated isotopic colonic transit time compared with the control group. We conclude that in healthy humans, 10-15 g ingestion or intracaecal infusion of lactulose produces a prolonged tonic contraction that may be involved in the laxative effect of lactulose.

Screening procedure for detection of stimulant laxatives and/or their metabolites in human urine using gas chromatography-mass spectrometry after enzymatic cleavage of conjugates and extractive methylation.

A gas chromatography-mass spectrometry (GC-MS)-based screening procedure was developed for the detection of stimulant laxatives and/or their metabolites in human urine after enzymatic cleavage of conjugates followed by extractive methylation. The part of the phase-transfer catalyst remaining in the organic phase was removed by solid-phase extraction on a diol phase. The compounds were separated by capillary GC and identified by computerized MS in the full scan mode. By use of mass chromatography with the ions m/z 305, 290, 335, 320, 365, 350, 311, 326, 271, and 346, the possible presence of stimulant laxatives and/or their metabolites could be indicated. The identity of positive signals in such mass chromatograms was confirmed by comparison of the peaks underlying full mass spectra with the reference spectra. This method allowed the detection of the diphenol laxatives bisacodyl, picosulfate, and phenolphthalein and of the anthraquinone laxatives contained in plant extracts and/or their metabolites in human urine samples. The overall recoveries of the stimulant laxatives and/or their metabolites ranged between 33% and 89% with a coefficient of variation of less than 15%, and the limits of detection ranged between 10 and 25 ng/mL (S/N 3) in the full scan mode. After ingestion of the lowest therapeutic dose of sodium picosulfate, its main metabolite, bisacodyl diphenol, was detectable in urine samples for 72 hours. After ingestion of the lowest therapeutic dose of a senna extract, the main metabolite of sennosides, rhein, was detectable in urine samples for 24 hours. This procedure is part of a systematic toxicological analysis procedure for acidic drugs and poisons with the modification of enzymatic cleavage of conjugates.

Soluble dietary fiber protects against cholesterol gallstone formation.

BACKGROUND: Epidemiological studies have suggested that soluble dietary fibers are hypocholesterolemic and may inhibit cholelithiasis. METHODS: Thirty prairie dogs were placed on a cholesterol-supplemented lithogenic diet. Ten animals received 5% psyllium (PSY) and 10 animals received 5% cellulose. After 6 weeks all gallbladders were inspected for stones; blood and bile were collected for analysis. RESULTS: Cholesterol stones were present in 8 of 10 of the control animals, in 6 of 10 of the cellulose group, and 3 of 10 of the PSY animals (P <0.05). Concentrations of cholesterol and chenodeoxycholic acid (CDCA) were significantly lower in the PSY group compared with controls (0.49 versus 0.88 mM and 4.2 versus 9.2 mM, respectively) leading to a significant reduction in the cholesterol saturation index (0.62 versus 1.2). CONCLUSIONS: A dietary soluble fiber (PSY) inhibits cholesterol stone formation by reducing the biliary cholesterol saturation index. This protective effect is associated with a selective decrease in biliary cholesterol and CDCA.

Review article: anthranoid laxatives and their potential carcinogenic effects.

Anthranoid laxatives are widely used laxatives of natural origin. Because of their chemical structure they are carried unabsorbed to the large bowel, where metabolism to the active aglycones takes place. These aglycones exert their laxative effect by damaging epithelial cells, which leads directly and indirectly to changes in absorption, secretion and motility. Damaged epithelial cells can be found as apoptotic bodies in the pigmented colonic mucosa, characteristic for pseudomelanosis coli. Pseudomelanosis coli is a condition caused by chronic (ab)use of anthranoid laxatives and has recently been associated with an increased risk of colorectal carcinoma. In vitro and animal studies have shown a potential role of anthranoid laxatives in both the initiation and promotion of tumorigenesis. Studies in humans have also suggested tumour promoting activities for these laxatives. Although the short-term use of these substances is generally safe, long-term use cannot be recommended.

Studies of aloe. VI. Cathartic effect of isobarbaloin.

The cathartic effect of isobarbaloin, a stereoisomer of barbaloin (compound principally responsible for the cathartic activity of Aloe), was examined in male rats by oral administration. Individual differences in sensitivity in the laxative activity of isobarbaloin and barbaloin was not found. The cathartic activity (ED50) of isobarbaloin in barbaloin positive rats was 19.2 mg/kg, nearly equal to that of barbaloin (19.5 mg/kg). Also, isobarbaloin administered orally was demonstrated to decompose to aloe-emodin-9-anthrone (active metabolite of barbaloin) as well as to barbaloin. Therefore, it is considered that the mechanism underlying the cathartic effect of isobarbaloin is the same as that of barbaloin.

[The mechanism of action of sennosides]. / Mécanisme d'action des sennosides.

A review of the recent progress in the study of the mode of action of the sennosides, the active constituents of the senna drug, is presented. An interaction between rhein anthrone, the active metabolite of the sennosides, and the immune cells of the colon is suggested as a base for laxative activity.

Cleavages of the O- and C-glucosyl bonds of anthrone and 10,10'-bianthrone derivatives by human intestinal bacteria.

A strictly anaerobic bacterium, Bifidobacterium sp. SEN, capable of hydrolyzing the O-glucosyl of sennosides was isolated from human feces. The bacterium stepwisely hydrolyzed sennoside B to sennidin B through sennidin-8-monoglucoside in PYF medium but not in GAM broth. Addition of D-glucose to PYF medium resulted in loss of the hydrolyzing activity in culture but addition of D-fructose did not affect the activity. Coculture of this bacterium with Peptostreptococcus intermedius led to rapid accumulation of rhein anthrone in the medium. Similarly, a bacterium, Eubacterium sp. BAR, capable of cleaving the C-glucosyl of barbaloin was isolated from human feces. This bacterium grew in PYF medium containing barbaloin and produced enzyme(s) that cleave(s) the C-glucosyl. The induction of the enzymes was completely inhibited in the presence of D-glucose. Nojirimycin inhibited the enzyme activity induced by barbaloin but it did not inhibit the bacterial growth in the presence of D-glucose.

Metabolism and pharmacokinetics of anthranoids.

Anthranoid derivatives are used all over the world as a treatment for constipation. These compounds are present in several drugs of plant origin, especially as O- or C-glycosides. Besides featuring different substituents, the aglycone might consist of an anthraquinone, an anthrone or a dianthrone. So far, detailed information concerning their metabolism and pharmacokinetic characteristics is available only in a few cases. The best characterized compounds are sennoside, a dianthrone O-glycoside present in senna leaves and senna pods, and its aglycone (rhein anthrone). After oral administration, sennoside is degraded only in the lower parts of the gastrointestinal tract, releasing its active metabolite rhein anthrone. Nowadays, this process is understood at the molecular level. A study with 14C-labelled rhein anthrone administered intracecally to rats, revealed that the compound is scarcely absorbed. Since on the contrary its anthraquinone equivalent is absorbed to a much larger extent, it is inferred that dianthrone- or anthrone-glycosides exhibit a lower systemic availability than anthraquinone O-glycosides.

The dependence of the in vitro fermentation of dietary fibre to short-chain fatty acids on the contents of soluble non-starch polysaccharides.

The fermentability of cellulose and dietary fibre in common clinical use (Inolaxol, Fiberform, Vi-Siblin, Lunelax, pectin) was measured as the in vitro production of short-chain fatty acids, lactate, and ammonia in 16.6% faecal homogenates from 18 healthy volunteers. The results were compared with the contents of soluble and insoluble non-starch polysaccharides as determined by the method of Englyst. The amounts of soluble non-starch polysaccharides in the fibre were closely associated with the mean productions of short-chain fatty acids after 6 h (R = 0.94, p < 0.002) and 24 h (R = 0.98, p < 0.0002) of incubation. The mean production of ammonia was inversely related to the soluble fraction of the fibre (after 6 h, R = -0.93, p < 0.003; after 24 h, R = -0.90, p < 0.006). These variables were not dependent on the insoluble fractions of the fibre. The in vitro fermentability differed considerably among the fibres: cellulose and Inolaxol (sterculia gum) were almost non-fermentable, Fiberform (wheat bran-based) was low-grade fermentable, Vi-Siblin and Lunelax (both ispaghula husk) were intermediately fermentable, and pectin was highly fermentable. These findings support that the water solubility determines the degree of fermentability of dietary fibre and thereby the corresponding bacterial assimilation of ammonia. In vitro measurements of short-chain fatty acid production in faecal homogenates may hence supplement commonly used methods to classify dietary fibre.