Comparison of rat and human intestinal perfusion models for assessing efficacy of oral rehydration solutions

The optimal composition for oral rehydration solutions remains controversial. Animal models have been used to assess the efficacy of new formulations but the relevance of these studies to the handling of oral rehydration solutions in human intestine during diarrhoeal disease states remains uncertain. Using steady state perfusion techniques we have compared water and solute transport from a variety of oral rehydration solutions in both the entire rat small intestine and in the human jejunum. Overall the pattern of water, sodium and glucose absorption was similar from the three oral rehydration solutions tested, indicating close parallelism between the two models despite the species and methodological differences. Although the relationship between the findings of these studies to the handling of oral rehydration solution in diarrhoeal disease states remains uncertain, we believe they do support the view that animal models may have a part to play in the preliminary screening of oral rehydration solutions before clinical trial.     

口服补液盐溶液对轻至中度脱水的疗效和安全性

目的:评价口服补液盐溶液治疗儿童呕吐和腹泻所致轻、中度脱水的疗效和安全性.方法:采用多中心、随机、对照的试验设计.试验组66例,对照组65例完成研究.试验组服用口服补液盐溶液(新配方),对照组服用口服补液盐Ⅱ(WHO推荐配方),用于迅速补充患儿因腹泻或呕吐所致的水和电解质的丢失(体液丢失量不超过5%～10%).结果:试验组和对照组改善体重减轻的疗效分别为100%和95.4%;改善皮肤弹性差等一般体征的疗效分别为97.0%和96.9%;改善循环系统体征的疗效分别为95.9%和100%;改善神经系统症状的疗效均为100%;改善症状总积分的疗效均为100%.2组之间所有症状和症状总积分比较差异均无显著性(P＞0.05).结论:新配方口服补液盐溶液治疗儿童呕吐和腹泻所致轻、中度脱水的疗效确切,可迅速补充水和电解质,不良反应少.     

双苓止泻口服液治疗小儿肠炎的疗效

我们于１９９９年１０月至２０００年１月对双苓止泻口服液(太极集团涪陵制药厂生产,批号:９９０７１５)治疗小儿肠炎的疗效进行了临床观察,现将观察结果总结如下:     

Assessment of water and solute absorption from experimental hypotonic and established oral rehydration solutions in secreting rat intestine

Water and solute absorption from three experimental hypotonic oral rehydration solutions (HYPO-ORS; sodium 45, 60 and 75 mmol/L, glucose 90 mmol/L), the World Health Organization recommended ORS (WHO-ORS; sodium 90 mmol/L, glucose 111 mmol/L), and the British National Formulary recommended ORS (BNF-ORS; sodium 35 mmol/L, glucose 200 mmol/L), have been assessed by perfusion studies in cholera toxin-induced secreting rat intestine. Net water absorption was greatest from the most hypotonic solution (HYPO-45; P less than 0.05). UK-ORS prevented net water secretion and WHO-ORS promoted moderate net water absorption. Net sodium secretion was seen with all solutions but was least from WHO-ORS and greatest with BNF-ORS (P less than 0.01). Glucose absorption was similar from BNF-ORS, WHO-ORS and HYPO-45 and in each case was greater than glucose absorption from HYPO-60 and HYPO-75 (P less than 0.05). These results suggest that net water and sodium absorption from ORS may be enhanced if osmolality is reduced by decreasing the glucose content.     

Studies of water movement across the gut using oral rehydration solutions in a rat perfusion model

Aim: To measure water influx and efflux, as well as net water, sodium and potassium absorption from a range of oral rehydration solutions (ORS) in which the glucose content had been partially replaced with the amino acid leucine or with food supplements. Methods: A series of in vivo steady-state perfusion studies in normal rat intestine. The oral rehydration solutions contained 60 or 90 mmol/L of sodium. The reference solution used was the World Health Organization (WHO) formula. Results: There was a significant negative correlation between the oral rehydration solution osmolality and net water absorption (r=–0.722, P < 0.05). The highest net water absorption occurred using comminuted chicken supplemented oral rehydration solution containing 60 mmol/L sodium (P < 0.001). This oral rehydration solution also showed a significant increase in the rate of influx of water (P < 0.05) in comparison with the WHO formula containing 60 mmol/L sodium. Conclusion: This work provides further evidence that food-based oral rehydration solutions, including non-vegetable sources, may have a useful role to play in the management of patients with acute diarrhoea.     

Excipients enhance intestinal absorption of ganciclovir by P-gp inhibition: assessed in vitro by everted gut sac and in situ by improved intestinal perfusion

In rats we examined the effects of some common excipients on the intestinal absorption of ganciclovir (GCV), a BCS-III drug and substrate of P-gp, by assessing its in vitro transfer from mucosa to serosa and in situ transepithelial permeation. In vitro, all selected excipients (concentration range 0.1-1% [w/v]) could increase the transport amount of GCV in the everted gut sac model. Whereas enhancement by F-68 demonstrated regional differences like verapamil, PEG-400, Tween-80 and EL-35 exhibited no regional differences. In situ studies were performed by an improved perfusion model, single-pass perfusion with whole small intestine, to determine more accurately the permeability of lipophobic compounds. The permeability of GCV was significantly increased by all excipients. The effects of EL-35 and F-68 were dose-dependent but those of PEG-400 and Tween-80 were not. The results suggest that enhancements of intestinal absorption of GCV by these excipients are probably due to inhibition of P-gp-mediated drug efflux. It could be deduced from their different properties that both blocking binding sites of P-gp and altering membrane fluidity were involved in their P-gp-inhibition. The former mechanism might be involved for F-68, while the latter one might account for the effects of PEG-400, Tween-80 and EL-35.     

高效液相色谱法同时测定大鼠肠灌流液中L-肌肽和酚红的浓度

目的建立HPLC方法同时测定大鼠肠灌流液中L-肌肽和酚红的质量浓度。方法采用KromasilNH2色谱柱（200mm×4．6mm,5μm）,流动相为乙腈-磷酸盐缓冲液（1mmol．L^-1磷酸二氢钾,0．2mmol．L^-1磷酸氢二钾）（体积比50：50）,流速1．0mL·min^-1,柱温35℃,检测波长210nm,进样量20μL。结果L-肌肽和酚红分别在质量浓度22．6～158mg·L^-1（r=0．9998）、14．2～99．4mg·L^-1（r=0．9991）与峰面积线性关系良好,2种成分回收率分别为99．1％、98．8％,RSD均〈3％。结论所用方法准确、简便、专属性好,可准确快速测定大鼠肠灌流液中L-肌肽和酚红的含量;L-肌肽的摄取过程依赖于H^＋电位梯度。     

Permeability of human intestinal mucosa using a continuous flow-through perfusion system

Continued interest in in vitro methods for performing bioavailability/bioequivalence (BA/BE) studies for drug registration purposes, prompted us to investigate the suitability of a continuous flow-through perfusion system to determine diffusion of a wide variety of permeants, through human intestinal mucosa. Permeability of fresh and frozen intestinal mucosa towards water, 17beta-estradiol, sumatriptan, arecoline and vasopressin was compared. Furthermore, diffusion studies of water, sumatriptan, arecoline, arecaidine, estradiol, cyclosporin and vasopressin across frozen/thawed intestinal mucosa specimens (-85 degrees C) were performed. No statistically significant differences between the flux values of the five compounds tested across fresh and frozen intestinal tissue, were found. Furthermore, it was demonstrated that the flux rates of the various compounds across these tissues decreased with increasing molecular size. However, the flux rates across frozen intestinal mucosa for compounds with molecular weights >300 Da, were low. Flux rates for the compounds studied across frozen/thawed human vaginal and buccal mucosa were 36-160% higher than those across frozen intestinal mucosa. We concluded that the continuous flow-through perfusion system used shows promise as an in vitro method for permeability determination through intestinal mucosa. However, other human mucosa e.g. vaginal mucosa, may have to be considered as alternatives to intestinal mucosa if therapeutic agents with molecular weights >500 Da are to be compared for in vitro BA/BE purposes, and further studies in this respect are warranted.     

反相高效液相色谱法同时测定大鼠单向肠灌流液中补骨脂素和异补骨脂素

目的建立同时测定大鼠在体单向肠灌流液中补骨脂素和异补骨脂素的RP-HPLC方法。方法采用RP-HPLC法,色谱柱为ShimadzuC18（250mm×4.6mm,5μm）,流动相为甲醇-水（55∶45）,柱温：30℃,流速：1.0mL·min^-1,检测波长：245nm。结果补骨脂素、异补骨脂素分别在0.8236～32.94μg·mL^-1、1.140～45.60μg·mL^-1与峰面积呈良好线性关系,相关系数r均为0.9999。其平均回收率分别为98.98%、99.59%,RSD均〈3%。结论该法操作简便、结果准确、灵敏度高,可同时测定大鼠单向肠灌流液中补骨脂素和异补骨脂素。     

Toxicology digital sources produced and available in the United Kingdom (UK)

This paper looks at toxicology digital sources produced and available in the United Kingdom (UK). It includes products aggregating a number of sources, which may include information from other countries, and for the convenience of the reader and information seeker these have also been described.     

Fluoroquinolone (ciprofloxacin) secretion by human intestinal epithelial (Caco-2) cells

1. Human intestinal epithelial Caco-2 cells were used to investigate the mechanistic basis of transepithelial secretion of the fluoroquinolone antibiotic ciprofloxacin.
2. Net secretion and cellular uptake of ciprofloxacin (at 0.1 mM) were not subject to competitive inhibition by sulphate, thiosulphate, oxalate, succinate and para-amino hippurate, probenecid (10 mM), taurocholate (100 μM) or bromosulphophthalein (100 μM). Similarly tetraethylammonium and N-′methylnicotinamide (10 mM) were without effect.
3. Net secretion of ciprofloxacin was inhibited by the organic exchange inhibitor 4,4′-diisothiocyanostilbene-2-2′-disulphonic acid (DIDS, 400 μM).
4. Net secretion of ciprofloxacin was partially inhibited by 100 μM verapamil, whilst net secretion of the P-glycoprotein substrate vinblastine was totally abolished under these conditions. Ciprofloxacin secretion was unaltered after preincubation of cells with two anti-P-glycoprotein antibodies (UIC2 and MRK16), which both significantly reduced secretory vinblastine flux (measured in the same cell batch). Ciprofloxacin (3 mM) failed to inhibit vinblastine net secretion in Caco-2 epithelia, and was not itself secreted by the P-glycoprotein expressing and vinblastine secreting dog kidney cell line, MDCK.
5. Net secretion and cellular uptake of ciprofloxacin (at 0.1 mM) were not subject to alterations of either cytosolic or medium pH, or dependent on the presence of medium Na⁺, Cl⁻ or K⁺ in the bathing media.
6. The substrate specificity of the ciprofloxacin secretory transport in Caco-2 epithelia is distinct from both the renal organic anion and cation transport. A role for P-glycoprotein in ciprofloxacin secretion may also be excluded. A novel transport mechanism, sensitive to both DIDS and verapamil mediates secretion of ciprofloxacin by human intestinal Caco-2 epithelia.

     

Transesterification of p-hydroxybenzoate esters (parabens) by human intestinal (Caco-2) cells

p-Hydroxybenzoate ester (paraben) preservatives are used in numerous orally administered products. The recognized route of metabolism for parabens is hydrolysis to p-hydroxybenzoic acid followed by conjugation and excretion. However, in the presence of alcohols, a presystemic transesterification pathway not previously reported for the human intestine can occur. Using human intestinal (Caco-2) cells, it was observed that hydrolysis of parabens to p-hydroxybenzoic acid is reduced markedly by ethanol concentrations that can occur in the human intestine, 0.25-0.5% (v/v). Ethanol concentrations of 1.0-2.5% (v/v) were optimal for transesterification to ethylparaben in Caco-2 cell homogenates. The kinetics of the transesterification reaction with regard to ethanol concentration (0-20%), time, pH (3-9), protein concentration (1-5 mg ml-1) and substrate concentration (6.25-200 microM) as well as the effects of different alcohols were studied. The Km and Vmax values for transesterification with ethanol for methyl, propyl, butyl, heptyl and octyl parabens were 449.7, 165.7, 86.1, 24.2 and 45.9 microM and 114.4, 37.5, 19.5, 7.5 and 7.6 micromol h-1 mg-1 Caco-2 cell protein, respectively. The Vmax values for transesterification of methylparaben with ethanol, propan-1-ol, butan-1-ol were 114.4, 5.1 and 4.9 micromol h-1 mg-1, respectively. Collectively, the kinetic data demonstrate that the enzyme responsible for the transesterification reaction has a preference for short-chain esters and represents the first report of transesterification in human intestinal cells. An implication of this mechanism is that alcohol-containing in vitro biosystems or protocols for the study of parabens disposition could generate transesterified artefacts. The clinical or toxicological implication is that, following co-ingestion of ester compounds with ethanol, transesterification could provide the basis for a previously unrecognized drug-alcohol interaction.     

Efficacy of oral cobalamin (vitamin B12) therapy

Importance of the field: Cobalamin (vitamin B12) deficiency is particularly common in the elderly (>?15%). Management of cobalamin deficiency with cobalamin injections is well codified at present, but new routes of cobalamin administration (oral and nasal) are being studied, especially oral cobalamin therapy for food-cobalamin malabsorption.

Areas covered in this review: The objective of this review is to evaluate the efficacy of oral cobalamin treatment in elderly patients. To reach this objective, PubMed data were systematically searched for English and French articles published from January 1990 to July 2008. Data from our research group on cobalamin deficiency (Groupe d’Etude des CAREnce vitamine B12 – CARE B12) were also analyzed.

What the reader will gain: Three prospective randomized studies, a systematic review by the Cochrane group and five prospective cohort studies were found and provide evidence that oral cobalamin treatment may adequately treat cobalamin deficiency. The efficacy was particularly highlighted when looking at the marked improvement in serum vitamin B12 levels and hematological parameters, for example hemoglobin level, mean erythrocyte cell volume and reticulocyte count. The effect of oral cobalamin treatment in patients presenting with severe neurological manifestations has not yet been adequately documented. Oral cobalamin treatment avoids the discomfort, inconvenience and cost of monthly injections.

Take home message: Our experience and the present analysis support the use of oral cobalamin therapy in clinical practice.     

Efficacy of a mucoadhesive patch compared with an oral solution for treatment of aphthous stomatitis

OBJECTIVE: The purpose of this study was to evaluate the efficacy and tolerability of a mucoadhesive patch compared with a pain-relieving oral solution for the treatment of aphthous stomatitis. METHODS: Patients with active aphthous stomatitis were randomly treated either once a day with a mucoadhesive patch containing citrus oil and magnesium salts (n = 26) or three times a day with an oral solution containing benzocaine and compound benzoin tincture (n = 22). All patients were instructed to apply the medication until pain had resolved, and completed a questionnaire detailing multiple clinical parameters followed by an evaluation of the treatment. RESULTS: The mucoadhesive patch was found to be more effective than the oral solution in terms of healing time (mean +/- SD: 36.0 +/- 22.8 hours vs 134.7 +/- 57.7, p < 0.001) and pain intensity after 12 and 24 hours (3.7 +/- 2.8 vs 6.3 +/- 2.6, p = 0.003, and 2.3 +/- 2.7 vs 5.7 +/- 2.5, p < 0.001, respectively). Local adverse effects 1 hour after treatment were significantly (p < 0.01) less frequent among the mucoadhesive patch patients compared with the oral solution patients. CONCLUSIONS: The mucoadhesive patch was found to be significantly more effective and better tolerated than the oral solution in the treatment of aphthous stomatitis.     

Placental transfer and DNA binding of benzo(a)pyrene in human placental perfusion

Benzo(a)pyrene (BP) is the best studied polycyclic aromatic hydrocarbon, classified as carcinogenic to humans. The carcinogenic metabolite, benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE), binds covalently to DNA. The key enzyme in this metabolic reaction is CYP1A1, which has also been found in placenta and human trophoblastic cells. By using human placental perfusion we confirmed that BP added to the maternal circulation in concentrations of 0.1 and 1 μM reaches fetal compartment but somewhat slower than the freely diffusible reference substance antipyrine. A well-known P-glycoprotein (ABCB1/P-gp) antagonist verapamil did not affect the transfer more than it did in the case of antipyrine, indicating that ABCB1/P-gp does not have a role in BP transfer. In one of the two placentas perfused for 6 h with the higher concentration of BP (1 μM) BPDE specific DNA adducts were found in placental tissue after the perfusion, but not before. The ability of human trophoblastic cells to activate BP to BPDE–DNA adducts was confirmed in human trophoblastic BeWo cells. This study shows that maternal exposure to BP leads to the exposure of the fetus to BP and/or its metabolites and that placenta itself can activate BP to DNA adducts.