[Assessment of the state of the small intestine microbiota in children on a long-term dairy-free diet].

An overview of recent outcomes of studies indicates an imbalance in the diet of children. Quantitative and qualitative malnutrition of children is the basis of a number of childhood diseases. The aim of the research was to study the prevalence of small intestine bacterial overgrowth syndrome (SIBO) in children on a long-term dairy-free diet. Material and methods. 40 children aged 7-11 years following a long-term dairy-free diet (average 3 years and 5 months, from 0.5 to 6.3 years) were examined (main group). 30 children who did not follow restrictive diets were consisted control group. In all children, SIBO was determined using a hydrogen breath test with a load of lactulose using a digital analyzer of exhaled hydrogen. Results. The proportion of children with intolerance to dairy products was 32.5%: 10.0% with allergy to cow's milk proteins, and 22.5% with lactose intolerance. 27.5% children followed a dairy-free diet according to an unjustified prescription by physician. 30.0% of children did not consume dairy products because of their unwillingness. 10.0% of children did not consume dairy products due to the unwillingness of their parents. An imbalance in the microbiota of the small intestine during the hydrogen breath test with lactulose loading was detected in 55.0% of children following a long-term dairy-free diet. 22.5% of children complained of recurrent abdominal pain, diarrhea was determined in 10.0%, constipation - in 7.5%, nausea - in 10.0%. In the control group, the SIBO during the hydrogen breath test with lactulose loading was found in 20.0%. Periodic abdominal pain was determined in 10.0%, nausea - in 6.7%, diarrhea - in 10.0%, constipation - in 3.3% children. Conclusion. Thus, among children of primary school age who follow a long-term dairyfree diet, SIBO is significantly more often recorded relative to children who are on a traditional type of diet.

Intestinal metabolism and absorption mechanism of multi-components in Gaultheria leucocarpa var. yunnanensis - An assessment using in situ and in vitro models, comparing gut segments in pathological with physiological conditions.

ETHNOPHARMACOLOGICAL RELEVANCE: Dianbaizhu (Gaultheria leucocarpa var. yunnanensis) as a Chinese folk medicine exerts significant treatment effects on rheumatoid arthritis (RA) with a long historical time. Our previous reports showed that the anti-rheumatic arthritis fraction (ARF) extracted and enriched from Dianbaizhu possessed good druggability, which was better than its single active ingredients. However, the intestinal transport characteristics and mechanism of ARF have not been elucidated to date. AIM OF THE STUDY: In order to illustrate the role of active ingredients of ARF in alleviating RA and promoting the development of dosage forms, the intestinal metabolism, absorption properties and mechanism of ARF in vitro and in situ models were investigated. MATERIALS AND METHODS: Firstly, after incubating with 4 intestinal segments (duodenum, jejunum, ileum, and colon), 7 key components in ARF, including MATG-B, (+)-catechin, MSTG-A, Gaultherin, chlorogenic acid, quercetin, and kaempferol were quantitatively analyzed by a high-performance liquid chromatography (HPLC). Secondly, combining the physiological and pathological rats, the in situ single-pass intestinal perfusion and in vitro everted gut sacs of rats were performed to investigate the absorption features and transport mechanisms of ARF using HPLC and HPLC-Q-TOF-MS/MS. Subsequently, in situ studies were employed to determine the effect of P-glycoprotein (P-gp) inhibitor (verapamil) on the transport characteristics of ARF in RA model rats. RESULTS: Comparing the absorption parameters of ARF incubated in different intestinal segments, data showed that the absorption of ARF in the small intestine was significantly stronger than that of the colon (P < 0.01). The number of characterized prototype components was subjected to the incubation time, drug concentration and rat body condition, but not the intestinal segments. There were no significant differences in the number of metabolites among different intestinal segments, administration concentrations and incubation time. The best small intestinal absorption site of ARF was duodenum and ileum in normal and model rats, respectively. The Peff values of 7 index compounds were all higher than 0.2 × 10-4cm/s, and the Fa values of 7 index compounds were all greater than 20% in the in situ perfusion investigation. The results showed that MSTG-B, MSTG-A and Gaultherin were likely to be substrates of P-gp as verapamil significantly enhanced their Peff and Ka values, while other ingredients were not P-gp substrates. CONCLUSIONS: The intestinal membrane permeability of ARF was good. Its intestinal absorption mechanisms mainly involved active transportation processes and passive diffusion. Besides, this report provided data support and basis for clinical development, bioavailability improvement and formulation design.

Assessment of small-intestine permeability in healthy Nigerian children is altered by urinary volume and voiding status.

OBJECTIVE: This study aimed to uncover the effect of voided urinary volume on small intestine permeability ratios in healthy children. METHODS: We assessed small intestine permeability in 155 apparently healthy children, aged 3-5 years old, without any visible symptoms of disease, in a rural, malaria-endemic setting in Nigeria, using a multi-sugar test solution, comprising lactulose, sucrose, mannitol, and rhamnose. Children were categorized into low urinary volume (LV) and high urinary volume (HV), based on the volume of urine voided per kg body weight per hour. LV children voided less than 25th percentile of the total population, while HV children voided greater than 75th percentile of the total population. Urinary volume excreted over a 90-minute period after administration of the test solution was measured, and differences in sugar ratios were compared between children with high (HV) and low urinary volumes (LV), as well as between children who voided (VC) or who were not able to void (NVC) before administration of the test solution. RESULTS: Urinary mannitol and rhamnose recovery were 44% (p = 0.002) and 77% (p<0.001) higher in HV children compared to LV children respectively, while urinary lactulose recovery was 34% lower (p = 0.071). There was no difference in urinary sucrose recovery between groups (p = 0.74). Lactulose-mannitol ratio, lactulose-rhamnose ratio and sucrose-rhamnose ratio were all significantly higher in children in the LV group compared to children in the HV group (p<0.001). In a multiple regression analysis, urinary volume and voiding status combined, explained 13%, 23% and 7% of the variation observed in lactulose-mannitol, lactulose-rhamnose and sucrose-rhamnose ratios, respectively. CONCLUSION: Sugar permeability ratios vary significantly with total urinary volume in multi-sugar small-intestine permeability tests. Voiding status before sugar administration appears to influence lactulose recovery, lactulose-rhamnose and sucrose-rhamnose ratios independently of total urinary volume. Evidence from this study suggests the need to take urinary volume into account when conducting multi-sugar small-intestine permeability tests.

A Medium-Throughput System for In Vitro Oxidative Stress Assessment in IPEC-J2 Cells.

The feed industry continuously seeks new molecules with antioxidant capacity since oxidative stress plays a key role in intestinal health. To improve screening of new antioxidants, this study aims to set up an assay to assess oxidative stress in the porcine small intestinal epithelial cell line IPEC-J2 using plate-reader-based analysis of fluorescence. Two oxidants, H2O2 and menadione, were tested at 1, 2 and 3 mM and 100, 200 and 300 µM, respectively. Trolox (2 mM) was used as the reference antioxidant and the probe CM-H2DCFDA was used to indicate intracellular oxidative stress. Cell culture, reactive oxygen species (ROS) production and assessment conditions were optimized to detect a significant ROS accumulation that could be counteracted by pre-incubation with trolox. Menadione (200 µM) reproducibly increased ROS levels, H2O2 failed to do so. Trolox significantly decreased intracellular ROS levels in menadione (200 µM)-exposed cells in a consistent way. The system was further used to screen different concentrations of the commercially available antioxidant ELIFE®. Concentrations between 100 and 200 ppm protected best against intracellular ROS accumulation. In conclusion, the combination of CM-H2DCFDA fluorescence analysis by a plate-reader, trolox as a reference antioxidant and 200 µM of menadione as a stressor agent, provides a replicable and reliable medium-throughput setup for the evaluation of intracellular oxidative stress in IPEC-J2 cells.

Assessment of pharmacokinetic variations of capecitabine after multiple administration in rats: a physiologically based pharmacokinetic model.

PURPOSE: Capecitabine is a prodrug of 5-fluorouracil (5-FU) used for the treatment of colorectal cancer, with a two-week course of administration. However, the variance in plasma concentration and metabolic enzyme activities after multiple administration of capecitabine and its metabolites is unknown. The aim of this study was to identify the variance and predict the plasma concentration profile of capecitabine and its metabolites, using metabolic enzyme activities, to develop a more effective and safer medication. METHODS: Rats orally received 180 mg/kg of capecitabine once a day for two weeks. Blood samples were collected nine times, and plasma concentration was measured on day 1, 7, and 14. The liver and small intestine were removed after blood sampling and were used in vitro to evaluate metabolic enzyme activities of carboxylesterase, cytidine deaminase, and thymidine phosphorylase. A physiologically based pharmacokinetic (PBPK) model was developed using in vitro results. RESULTS: Area under the plasma concentration-time curve from 0 h to infinity of 5-FU on day 7 and day 14 was significantly lower than that on day 1. Intrinsic clearance of thymidine phosphorylase in the liver on day 7 and day 14 was 1.4 and 1.3 times lower than that on day 1, respectively. The PBPK model described the observed plasma concentration of capecitabine and its metabolites. CONCLUSION: The decreased plasma concentration of capecitabine was caused by decreased metabolic enzyme activity. Efficacy can be improved by dose adjustment of capecitabine based on metabolic enzyme activities, using the PBPK model.

Evaluation of neutral oral contrast agents for assessment of the small bowel at abdominal staging CT.

BACKGROUND: Although neutral oral contrast agents are widely in use, a consensus regarding a standardized protocol in abdominal staging CT does not exist. PURPOSE: To test the null hypothesis that there is no quantitative or qualitative difference between water and mannitol for evaluation of the small bowel at abdominal staging CT. MATERIAL AND METHODS: 180 patients prospectively underwent abdominal staging CT with oral administration of either 1 liter mannitol solution (n = 88) or water (n = 92). Intestinal distension was measured in 6 different segments of the small intestine. In addition, two radiologists separately evaluated diagnostic image quality with regards to luminal distension (three-point scale) in each segment and the possibility to rule out a possible underlying pathology. Quantitative and qualitative results were compared (Mann-Whitney test). RESULTS: Quantitatively, intestinal distension was comparable in all segments (p>0.05), except for the horizontal duodenum (p = 0.019). The mean luminal diameter over all intestinal segments was 19.0 mm (18.1-19.9 mm) for the water group and 18.4 mm (17.5-19.2 mm) for the mannitol group, respectively. Qualitatively, ratings were comparable for the first three segments, while distal segments were rated better using mannitol. Side effects were only observed using mannitol (n = 26; 29.5%). CONCLUSIONS: Orally administered water and mannitol solution for evaluation of the small bowel at abdominal staging CT in clinical routine resulted in comparable results for the quantitative, but not for the qualitative analysis. Looking more differentiated at the overall performance, water has advantages in terms of patient comfort, side effects and costs, and can therefore be regarded as noninferior to mannitol in this specific patient group.

Combining in vitro digestion model with cell culture model: Assessment of encapsulation and delivery of curcumin in milled starch particle stabilized Pickering emulsions.

To investigate the encapsulation and oral delivery efficiency of milled starch particles stabilized Pickering emulsions for lipophilic bioactive compounds, in vitro digestion model coupled with Caco-2 cells models were used. Physicochemical and biological properties of curcumin encapsulated Pickering emulsions were analyzed regarding to emulsion structure, curcumin retention, in vitro digestion, in vitro anti-proliferate ability and cellular uptake. Milled starch particles stabilized Pickering emulsion system was able to protect curcumin against harsh gastric conditions. Around 80% of the encapsulated curcumin was retained after 2 h of simulated gastric digestion. By being encapsulated in Pickering emulsion, the bioaccessibility of curcumin was increased from 11% for curcumin in bulk oil phase to 28% under simulated intestinal digestion process. The resulting curcumin-loaded micelle phase from digested emulsion exhibited significant anti-cancer ability and enhanced cellular uptake. This research provides an exploratory study on the possible future application of milled starch particles stabilized Pickering emulsions as nutraceutical delivery vehicles in the creation of novel functional foods.

Arsenic in Rice Bran Products: In Vitro Oral Bioaccessibility, Arsenic Transformation by Human Gut Microbiota, and Human Health Risk Assessment.

Despite rice consumption, rice bran as a byproduct of rice milling contains higher arsenic (As). The present study evaluated the metabolic potency of in vitro cultured human colon microbiota toward As from five rice bran products with 0.471-1.491 mg of As/kg. Arsenic bioaccessibility ranged from 52.8 to 78.8% in the gastric phase, and a 1.2-fold increase (66.0-95.8%) was observed upon the small intestinal phase. Subsequently, a significant decline of As bioaccessibility (11.3-63.6%) and a high methylation percentage of 18.5-79.8% were found in the colon phase. The predominant As species in the solid phase was always As(V) (49.6-63.4%), and As-thiolate complexes increased by 10% at the end of colon incubation. Human gut microbiota could induce As bioaccessibility lowering and As transformation in rice bran, which illustrated the importance of food-bound As metabolism in the human body. This will result in a better understanding of health implications associated with As exposures.

Oxfendazole kinetics in pigs: In vivo assessment of its pattern of accumulation in Ascaris suum.

Ascaris suum is a widespread parasitic nematode that causes infection in pigs with high prevalence rates. Oxfendazole (OFZ) is effective against A. suum when used at a single high oral dose of 30 mg/kg. The aim of this study was to assess the pattern of distribution/accumulation of OFZ and its metabolites, in bloodstream (plasma), mucosal tissue and contents from small and large intestine and adult specimens of A. suum collected from infected and treated pigs. The activity of glutathione-S-transferases (GSTs) in A. suum was also investigated. Infected pigs were orally treated with OFZ (30 mg/kg) and sacrificed at 0, 3, 6 and 12 h after treatment. Samples of blood, mucosa and contents from both small and large intestine as well as adult worms were obtained and processed for quantification of OFZ/metabolites by HPLC. OFZ was the main analyte measured in all of the evaluated matrixes. The highest drug concentrations were determined in small (AUC0-t 718.7 ±â€¯283.5 µg h/g) and large (399.6 ±â€¯110.5 µg h/g) intestinal content. Concentrations ranging from 1.35 to 2.60 µg/g (OFZ) were measured in adult A. suum. GSTs activity was higher after exposure to OFZ both in vivo and ex vivo. The data obtained here suggest that the pattern of OFZ accumulation in A. suum would be more related to the concentration achieved in the fluid and mucosa of the small intestine than in other tissues/fluids. It is expected that increments in the amount of drug attained in the tissues/fluids of parasite location will correlate with increased drug concentration within the target parasite, and therefore with the resultant treatment efficacy. The results are particularly relevant considering the potential of OFZ to be used for soil transmitted helminths (STH) control programs and the advantages of pigs as a model to assess drug treatment to be implemented in humans.

In vitro methods to assess drug precipitation in the fasted small intestine - a PEARRL review.

OBJECTIVES: Drug precipitation in vivo poses a significant challenge for the pharmaceutical industry. During the drug development process, the impact of drug supersaturation or precipitation on the in vivo behaviour of drug products is evaluated with in vitro techniques. This review focuses on the small and full scale in vitro methods to assess drug precipitation in the fasted small intestine. KEY FINDINGS: Many methods have been developed in an attempt to evaluate drug precipitation in the fasted state, with varying degrees of complexity and scale. In early stages of drug development, when drug quantities are typically limited, small-scale tests facilitate an early evaluation of the potential precipitation risk in vivo and allow rapid screening of prototype formulations. At later stages of formulation development, full-scale methods are necessary to predict the behaviour of formulations at clinically relevant doses. Multicompartment models allow the evaluation of drug precipitation after transfer from stomach to the upper small intestine. Optimisation of available biopharmaceutics tools for evaluating precipitation in the fasted small intestine is crucial for accelerating the development of novel breakthrough medicines and reducing the development costs. SUMMARY: Despite the progress from compendial quality control dissolution methods, further work is required to validate the usefulness of proposed setups and to increase their biorelevance, particularly in simulating the absorption of drug along the intestinal lumen. Coupling results from in vitro testing with physiologically based pharmacokinetic modelling holds significant promise and requires further evaluation.

Relationships between milk mid-IR predicted gastro-enteric methane production and the technical and financial performance of commercial dairy herds.

Considering economic and environmental issues is important in ensuring the sustainability of dairy farms. The objective of this study was to investigate univariate relationships between lactating dairy cow gastro-enteric methane (CH4) production predicted from milk mid-IR (MIR) spectra and technico-economic variables by the use of large scale and on-farm data. A total of 525 697 individual CH4 predictions from milk MIR spectra (MIR-CH4 (g/day)) of milk samples collected on 206 farms during the Walloon milk recording scheme were used to create a MIR-CH4 prediction for each herd and year (HYMIR-CH4). These predictions were merged with dairy herd accounting data. This allowed a simultaneous study of HYMIR-CH4 and 42 technical and economic variables for 1024 herd and year records from 2007 to 2014. Pearson correlation coefficients (r) were used to assess significant relationships (P<0.05). Low HYMIR-CH4 was significantly associated with, amongst others, lower fat and protein corrected milk (FPCM) yield (r=0.18), lower milk fat and protein content (r=0.38 and 0.33, respectively), lower quantity of milk produced from forages (r=0.12) and suboptimal reproduction and health performance (e.g. longer calving interval (r=-0.21) and higher culling rate (r=-0.15)). Concerning economic results, low HYMIR-CH4 was significantly associated with lower gross margin per cow (r=0.19) and per litre FPCM (r=0.09). To conclude, this study suggested that low lactating dairy cow gastro-enteric CH4 production tended to be associated with more extensive or suboptimal management practices, which could lead to lower profitability. The observed low correlations suggest complex interactions between variables due to the use of on-farm data with large variability in technical and management practices.

Assessment of bioavailable B vitamin content in food using in vitro digestibility assay and LC-MS SIDA.

Standardized analytical methods, where each B vitamin is extracted from a given sample individually using separate procedures, typically ensure that the extraction conditions provide the maximum recovery of each vitamin. However, in the human gastrointestinal tract (GIT), the extraction conditions are the same for all vitamins. Here, we present an analytically feasible extraction protocol that simulates conditions in the GIT and provides a measure of the content of bioavailable vitamins using LC-MS stable isotope dilution assay. The results show that the activities of both human gastric and duodenal juices were insufficient to liberate absorbable vitamers (AV) from pure cofactors. The use of an intestinal brush border membrane (IBBM) fraction derived from the mucosal tissue of porcine small intestine ensured at least 70% AV recovery. The rate of AV liberation, however, was strongly dependent on the cofactor, e.g., in the case of NADH, it was magnitudes higher than in the case of thiamine diphosphate. For some vitamins in some food matrices, the use of the IBBM fraction assay resulted in lower values for the content of AV than conventional vitamin determination methods. Conventional methods likely overestimate the actual bioavailability of some vitamins in these cases. Graphical abstract Assessment of bioavailable B vitamin content in food.

Assessment of in Vitro Digestibility of Dietary Carbohydrates Using Rat Small Intestinal Extract.

There are few studies on the assessment of digestibility of nondigestible carbohydrates, despite their increasingly important role in human health. In vitro digestibility of a range of dietary carbohydrates classified as digestible (maltose, sucrose, and lactose), well-recognized (lactulose, fructooligosaccharides (FOS), and two types of galactooligosaccharides (GOS) differing in the predominant glycosidic linkage), and potential (lactosucrose and GOS from lactulose, OsLu) prebiotics using a rat small intestinal extract (RSIE) under physiological conditions of temperature and pH is described. Recognized and potential prebiotics were highly resistant to RSIE digestion although partial hydrolysis at different extents was observed. FOS and lactulose were the most resistant to digestion, followed closely by OsLu and more distantly by both types of GOS and lactosucrose. In GOS, ß(1 → 6) linkages were more resistant to digestion than ß(1 → 4) bonds. The reported in vitro digestion model is a useful, simple, and cost-effective tool to evaluate the digestibility of dietary oligosaccharides.

Assessment of Bioequivalence of Weak Base Formulations Under Various Dosing Conditions Using Physiologically Based Pharmacokinetic Simulations in Virtual Populations. Case Examples: Ketoconazole and Posaconazole.

Postabsorptive factors which can affect systemic drug exposure are assumed to be dependent on the active pharmaceutical ingredient (API), and thus independent of formulation. In contrast, preabsorptive factors, for example, hypochlorhydria, might affect systemic exposure in both an API and a formulation-dependent way. The aim of this study was to evaluate whether the oral absorption of 2 poorly soluble, weakly basic APIs, ketoconazole (KETO) and posaconazole (POSA), would be equally sensitive to changes in dissolution rate under the following dosing conditions-coadministration with water, with food, with carbonated drinks, and in drug-induced hypochlorhydria. The systems-components of validated absorption and PBPK models for KETO and POSA were modified to simulate the above-mentioned clinical scenarios. Virtual bioequivalence studies were then carried out to investigate whether formulation effects on the plasma profile vary with the dosing conditions. The slow precipitation of KETO upon reaching the upper part of the small intestine renders its absorption more sensitive to the completeness of gastric dissolution and thus to the gastric environment than POSA, which is subject to extensive precipitation in response to a pH shift. The virtual bioequivalence studies showed that hypothetical test and reference formulations containing KETO would be bioequivalent only if the microenvironment in the stomach enables complete gastric dissolution. We conclude that physiologically based pharmacokinetic modeling and simulation has excellent potential to address issues close to bedside such as optimizing dosing conditions. By studying virtual populations adapted to various clinical situations, clinical strategies to reduce therapeutic failures can be identified.

Assessment of Small Intestinal Transit Times in Ulcerative Colitis and Crohn's Disease Patients with Different Disease Activity Using Video Capsule Endoscopy.

Small intestinal transit times (SITT) influence drug bioavailability. This study aimed to compare SITT in Crohn's disease and ulcerative colitis patients with non-inflammatory bowel disease (IBD) and to determine influence of disease activity on transit times, and in addition, to establish the utility of small bowel video capsule endoscopy (SB-VCE) in investigation of SITT in IBD patients. A retrospective review was performed on consecutive patients who had undergone SB-VCE at a university hospital out-patient clinic. In total, 125 non-IBD patients, 55 Crohn's disease patients, and 23 ulcerative colitis patients were included. SITT were calculated from the first duodenal image to the first cecal image. Disease activity was assessed based on endoscopy results and inflammatory markers (calprotectin, C-reactive protein, erythrocyte sedimentation rate). SITT were longer in ulcerative colitis patients compared to non-IBD patients (median 264 min vs. 216 min, p = 0.010). Patients with active Crohn's disease (n = 33) also displayed prolonged SITT compared to non-IBD patients (median 253 min vs 216 min, p = 0.017) and patients with quiescent Crohn's disease (n = 22) (p = 0.005). SITT can be prolonged in IBD patients depending on disease activity which may alter the drug release profiles and clinical response to colonic drug delivery systems. SB-VCE is a simple, non-invasive tool that can be utilized in pharmacokinetic studies to understand drug bioavailability in different patient groups. Moreover, this variability in transit times needs to be simulated in dissolution testing for in vitro in vivo correlations.

Ethanol oxidation and the inhibition by drugs in human liver, stomach and small intestine: Quantitative assessment with numerical organ modeling of alcohol dehydrogenase isozymes.

Alcohol dehydrogenase (ADH) is the principal enzyme responsible for metabolism of ethanol. Human ADH constitutes a complex isozyme family with striking variations in kinetic function and tissue distribution. Liver and gastrointestinal tract are the major sites for first-pass metabolism (FPM). Their relative contributions to alcohol FPM and degrees of the inhibitions by aspirin and its metabolite salicylate, acetaminophen and cimetidine remain controversial. To address this issue, mathematical organ modeling of ethanol-oxidizing activities in target tissues and that of the ethanol-drug interactions were constructed by linear combination of the corresponding numerical rate equations of tissue constituent ADH isozymes with the documented isozyme protein contents, kinetic parameters for ethanol oxidation and the drug inhibitions of ADH isozymes/allozymes that were determined in 0.1 M sodium phosphate at pH 7.5 and 25 °C containing 0.5 mM NAD(+). The organ simulations reveal that the ADH activities in mucosae of the stomach, duodenum and jejunum with ADH1C*1/*1 genotype are less than 1%, respectively, that of the ADH1B*1/*1-ADH1C*1/*1 liver at 1-200 mM ethanol, indicating that liver is major site of the FPM. The apparent hepatic KM and Vmax for ethanol oxidation are simulated to be 0.093 ± 0.019 mM and 4.0 ± 0.1 mmol/min, respectively. At 95% clearance in liver, the logarithmic average sinusoidal ethanol concentration is determined to be 0.80 mM in accordance with the flow-limited gradient perfusion model. The organ simulations indicate that higher therapeutic acetaminophen (0.5 mM) inhibits 16% of ADH1B*1/*1 hepatic ADH activity at 2-20 mM ethanol and that therapeutic salicylate (1.5 mM) inhibits 30-31% of the ADH1B*2/*2 activity, suggesting potential significant inhibitions of ethanol FPM in these allelotypes. The result provides systematic evaluations and predictions by computer simulation on potential ethanol FPM in target tissues and hepatic ethanol-drug interactions in the context of tissue ADH isozymes.

Assessment of the potential drug-drug interaction between carvedilol and clopidogrel mediated through intestinal P-glycoprotein.

The most widely prescribed oral antiplatelet agent, clopidogrel, shows high interindividual variability resulting in an increased risk of cardiovascular events in the patients with reduced platelet inhibition. The purpose of this study was to investigate the role of the P-glycoprotein (P-gp) efflux pump in limiting the intestinal permeability of clopidogrel and the effect of a ß-blocker, namely, carvedilol, on its intestinal transport. Effective permeabilities (Peff) of clopidogrel and carvedilol were investigated in the proximal jejunum and distal ileum of rats using an in situ intestinal perfusion model. Peff values of clopidogrel and carvedilol were found to be concentration dependent with decreased Peff values at the low perfusate concentrations. Coperfusion with the P-gp inhibitors verapamil (100 µM) and carvedilol (10 µM) significantly increased the Peff of clopidogrel in the jejunum (8.31±0.20 x 10-5 and 6.98±0.75 x 10-5 vs. 3.60±0.51 x 10-5, respectively) and ileum (9.08±2.19 x 10-5 and 8.35±1.58 x 10-5 vs. 3.85±0.15 x 10-5, respectively). However, at the highest concentration tested (30 µM), clopidogrel exhibited 3 and 1.4 times higher Peff than those of metoprolol, an FDA high permeability reference standard, in the jejunum and ileum, respectively. Overall, this study indicates that the efflux function appears not to have a significant impact on the in vivo intestinal absorption of clopidogrel due to the saturation of P-gp, suggesting no clinically relevant interaction between carvedilol and clopidogrel mediated through P-gp at intestinal level.

Low cost industrial production of coagulation factor IX bioencapsulated in lettuce cells for oral tolerance induction in hemophilia B.

Antibodies (inhibitors) developed by hemophilia B patients against coagulation factor IX (FIX) are challenging to eliminate because of anaphylaxis or nephrotic syndrome after continued infusion. To address this urgent unmet medical need, FIX fused with a transmucosal carrier (CTB) was produced in a commercial lettuce (Simpson Elite) cultivar using species specific chloroplast vectors regulated by endogenous psbA sequences. CTB-FIX (∼1 mg/g) in lyophilized cells was stable with proper folding, disulfide bonds and pentamer assembly when stored ∼2 years at ambient temperature. Feeding lettuce cells to hemophilia B mice delivered CTB-FIX efficiently to the gut immune system, induced LAP(+) regulatory T cells and suppressed inhibitor/IgE formation and anaphylaxis against FIX. Lyophilized cells enabled 10-fold dose escalation studies and successful induction of oral tolerance was observed in all tested doses. Induction of tolerance in such a broad dose range should enable oral delivery to patients of different age groups and diverse genetic background. Using Fraunhofer cGMP hydroponic system, ∼870 kg fresh or 43.5 kg dry weight can be harvested per 1000 ft(2) per annum yielding 24,000-36,000 doses for 20-kg pediatric patients, enabling first commercial development of an oral drug, addressing prohibitively expensive purification, cold storage/transportation and short shelf life of current protein drugs.

Toward oral delivery of biopharmaceuticals: an assessment of the gastrointestinal stability of 17 peptide drugs.

A major barrier to successful oral delivery of peptide and protein molecules is their inherent instability in the lumen of the gastrointestinal tract. The aim of this study was to determine the stability of 17 disparate peptide drugs (insulin, calcitonin, glucagon, secretin, somatostatin, desmopressin, oxytocin, [Arg(8)]-vasopressin, octreotide, ciclosporin, leuprolide, nafarelin, buserelin, histrelin, [d-Ser](4)-gonadorelin, deslorelin, and goserelin) in gastric and small intestinal fluids from both humans and pigs, and in simulated gastric and intestinal fluids. In human gastric fluid, the larger peptides including somatostatin, calcitonin, secretin, glucagon, and insulin were metabolized rapidly, while the smaller peptides showed good stability. In human small intestinal fluid, however, both small and large peptides degraded rapidly with the exception of the cyclic peptide ciclosporin and the disulfide-bridge containing peptides octreotide and desmopressin, which showed good stability. The stability of peptides in both simulated gastric fluid and pig gastric fluid correlated well with stability in human gastric fluid. However, it was not possible to establish such a correlation with the small intestinal fluids because of the rapid rate of peptide degradation. This work has identified the molecular features in the structure of a wide range of peptides that influence their stability in the environment of the gastrointestinal tract, which in turn will allow for better selection of peptide candidates for oral delivery.

[Toxicological assessment of nanostructured silica. The acute oral toxicity].

Nanostructured amorphous silica (SiO2) is one of the priorities of nanomaterials, exposing human to the ever-increasing scale as a component of food additives, drugs and cosmetic products. According to numerous publications SiO2 nanoparticles (NPs) possess deleterious effects on animal and human cells in vitro and also exhibit inhalation toxicity. However, the biological effects in vivo of silica NPs taken orally are studied insufficiently. This article represents the first section of this study which aim is identification of silica preparation as nanomaterial and estimating of acute toxicity after oral administration in the form of aqueous suspension. Studies of size and shape of the particles in aqueous suspension of silica used in the study by electron and atomic force microscopy, spectroaqustic analysis and dynamic laser light scattering showed that the test substance is a nanomaterial. Estimation of acute toxicity of an aqueous suspension of nanostructured silica with a single intragastric gavage to male BALB/C mice allowed to conclude that the test material has LD50 by the oral route greater than 10 000 mg/kg and consequently belongs to class IV (low danger agents). Pathological changes in jejunum and colon of mice in the acute experiment (at a dose of 10 000 mg/kg) haven't been identified. Thus SiO2 NPs possess low toxicity when administered in the gastrointestinal tract. The available data, however, do not exclude possible presence of adverse effects under their long-term administration by oral way.