Everted Sacs Research Articles

Oligomers Are Not the Limiting Factor in the Absorption of DL-2-Hydroxy-4-(methylthio)butanoic Acid in the Chicken Small Intestine

The methionine hydroxy analogue DL-2-hydroxy-4-(methylthio)butanoic acid (HMB) is commonly used as a supplemental source of methionine in commercial animal diets. The HMB free acid is an aqueous solution that contains 88% product in an equilibrium mixture of monomer, dimer, and polymeric compounds. The present study examines whether the presence of these nonmonomeric forms reduces the absorption of the hydroxy analogue in the chicken small intestine. In vivo and in vitro methodologies were used to compare the intestinal absorption of an HMB product containing mainly monomer (HMB-PCM) with commercial HMB. The results from the in vivo perfusion of the jejunum showed no significant differences between the 2 hydroxy analogue sources in monomer absorption from the intestinal lumen, tissue accumulation, or plasma concentration. The results also indicate that the nonmonomeric forms are hydrolyzed during perfusion. Moreover, monomer tissue accumulation in everted sacs showed no significant differences between substrates, either in the presence or in the absence of a H+-gradient; a higher value was observed in the jejunum and ileum in comparison with the duodenum. Similarly, serosal appearance in H+-gradient conditions did not differ significantly between substrates, and it showed the same regional profile as in tissue accumulation. Oligomer hydrolysis was confirmed in vitro without significant differences between segments. In conclusion, the presence of nonmonomeric forms is not a limiting factor in HMB absorption, apparently because of the hydrolytic capacity of intestinal mucosa, as confirmed by experiments in vivo and in vitro.

Modulatinn Intestinal Uptake of Atenolol Usinn Niosomes as Drun Permeation Enhancers

It is well established through the last decade that niosomes have potential applications as drug carriers either to improve drug permeation across membranes or targeting to specific tissues. Having a considerable ability to improve the permeability of drugs through lipoid membranes, niosomes have been utilized as carriers to enhance atenolol absorption from the gastrointestinal tract. Two methods have been adopted to prepare niosomes, the proniosome-derived method (A) and the conventional film hydration method (B). The products from the two methods were compared in terms of their morphology, vesicle size, drug encapsulation efficiency, in vitro drug release and enhancement effect on drug permeation across the intestinal membrane using an everted sac technique. Proniosome-derived niosomes were smoother and exhibited a smaller (5 μm) vesicle size compared to those prepared by conventional methods (12 μm). High encapsulation efficiencies of 98.6% and 93.4% were achieved by methods A and B, respectively. In vitro drug release has been significantly retarded from both types of niosomes. Comparing to pure drug, which dissolved completely in 15 min, only 8.9% and 9.9% of the entrapped drug was released in the same time period. The drug release kinetics showed non-Fickian (anomalous) behavior. Permeation through an everted intestinal sac showed a significant enhancement effect (more than 4 fold) for both types of niosomes compared to untrapped drug; however, the difference between the two types of niosomes was not significant

Characterization of secretory intestinal transport of the lactone form of CPT-11

It has been reported that a significant portion of the lactone form of 7-ethyl-10-(4-[1-piperidino]-1-piperidino)-carbonyloxy-camptothecin (CPT-11) is excreted into the gastrointestinal lumen via the intestinal membrane and that carboxylesterase activity, which converts CPT-11 to SN-38, was detected in the intestine. It is possible that a reduction in the excretion of CPT-11 lactone into the gastrointestinal lumen induces the gastrointestinal toxicity. The purpose of this study was to investigate the characteristics of transporter(s) that contribute to the jejunal efflux of the lactone form of CPT-11. The serosal-to-mucosal permeation rate of CPT-11 lactone was investigated in everted sac studies. The secretory transport required metabolic energy and was diminished by sulfobromophthalein (BSP) and 1-naphthol, inhibitors of the ME3277 transport system. However, inhibitors of breast cancer resistance protein (Bcrp), multidrug resistance-associated protein 2 (Mrp2) and P-glycoprotein (P-gp) did not affect the secretion of CPT-11 lactone. The results suggest that a specific transport system, which is identical to the ME3277 transport system, plays a major role in the secretion of CPT-11 lactone.

Ethyl α-D-glucoside was absorbed in small intestine and excreted in urine as intact form

Objective Ethyl α-D-glucoside (α-EG) is a peculiar component in sake. We investigated how α-EG was absorbed, hydrolyzed, and excreted in urine when it was ingested orally by rats. Methods Hydrolyzing activity for α-EG was determined by incubating it with crude enzyme solutions prepared from several rat organs, and absorption activity for α-EG was determined by incubating rat small intestinal everted sac in sodium or potassium Krebs-Ringer buffer that contained α-EG. α-EG solution was fed to rats, and urine volume and plasma α-EG, glucose and insulin and urinary α-EG were determined. Results α-EG was hydrolyzed by crude enzyme solutions prepared from rat small intestinal mucosa and kidney, and these hydrolyzing activities were lower than those for maltose. α-EG absorbed into everted rat intestinal sacs in potassium Krebs-Ringer buffer reduced almost completely compared with that in sodium Krebs-Ringer buffer. When α-EG was ingested orally by rats, it was absorbed into the bloodstream and more than 60% was excreted in urine, and urine volume increased. Conclusions In rats, α-EG was absorbed in small intestine and excreted intact in urine without affecting blood glucose and insulin and thus was a diuretic, insulin-independent, and low-nutritive glucoside that could be safely applicable to food.

Enteric Excretion of Baicalein, a Flavone of Scutellariae Radix, via Glucuronidation in Rat: Involvement of Multidrug Resistance-Associated Protein 2

Baicalin (BG) and its aglycone, baicalein (B), are strong antioxidants and have various pharmacological actions. The purpose of this study was to evaluate efflux of BG from rat intestinal mucosal cell following glucuronidation of B absorbed after oral administration of B. The absorption and excretion of BG and B were evaluated in rats using the in situ jejunal loop technique and in vitro jejunal everted sac experiments. BG and B levels were determined by high-performance liquid chromatography with electro-chemical detection to ensure selectivity and high sensitivity. A large amount (30.4% recovery) of BG, but no B, was detected in the intestinal lumens of germ-free rats 4 h after oral administration of B (12.1 mg/kg), in comparison with a substantial recovery (55.1%) of unabsorbed BG 4 h after its administration. During the in situ rat jejunal loop absorption experiment, B disappeared rapidly, and 8% of the lost B was excreted into the loop as BG 20 min after infusing 0.1 mM B. In an in vitro absorption experiment using everted rat jejunal sac, BG also appeared outside the sac, accompanied by the disappearance of B from the outer (mucosal) side. However, very little of B was transferred to the inner (serosal) side of the sac, and only a trace of BG was detected inside the sac. Thus, in both the loop and the everted sac systems, the efflux of BG from the mucosal surface was saturated with the concentration of B added. Moreover, the efflux rate of BG in the everted jejunal sac from Eisai hyperbilirubinemic rat (EHBR) was significantly lower by 56.4% than that from Sprague-Dawley rat. These results indicate that, in rat, a large proportion of any B absorbed is retained, transformed into BG within the intestinal mucosal cells, and coordinately excreted through multidrug resistance-associated protein 2 (MRP2) into the intestinal lumen.

Everted Intestinal Sacs As In vitro Model For Assessing Absorptivity Of L- Histidine Under The Effect Of Aspirine And Gum Acacia In Male Rats

The purpose of this study was to characterize intestinal permeability changes over a range of physiologically relevant intestinal injury. The experiments were performed in 80 rats subdivided into four groups as aspirin (400 mg/kg b.w.), gum Acacia ( 1g./day) and aspirin with gum Acacia groups for 21 days compared with control group. Relative reabsorption of L- Histidine was greater(p<0.001) in the aspirin in 10 min of incubation compared with that of the control rats. In aspirin in combination with gum Acacia, the relative reabsorption were significantly (p<0.001) decrease in 10, 20 and 30 min. of incubation compared with that of the control rats. Moreover, the relative reabsorption of L-histidine was significantly (p<0.01) reduced by the aspirin at 45 min of time of the incubation buffer compared with that of the control. However, gum acacia treatment was increased at 10 min (p<0.01) ,30 min (p<0.01) and 45 min (p<0.001) respectively compared with that of the control rats. Relative reabsorption of L-histidine record a nonsignificant increase of aspirin at 20 min and 30 min of incubation compared with that of the control. Gum and aspirin with gum at 20min and 45min of incubation resulted an increase and decrease in relative reabsorption of L-histidine respectively compared with that of the control. Aspirin and aspirin in combination with gum acacia treatment increased body, intestinal weights and mucosal total protein significantly with percent changes ranged from 8% to 40% compared with that of the control. On the other hand, gum treatment decreased body, intestinal weights and mucosal total protein significantly with percent changes ranged from 8% to 35% compared with that of the control. These results demonstrated that L-histidine is actively taken up by a gum Acacia system in intestinal everted sac mechanism of rat with energy supplied by glucose and Na + in incubation buffer. Moreover, aspirin system had an inhibitory effect on L-histidine uptake in 45 min of incubation , indicating the saturation by L-histidine in first ten minuts of incubation. Also, these results provide evidence the uptake of L-histidine into rat intestine was not reduced at all by the treatment of aspirin. These results suggest that the uptake of L-histidine by intestinal everted sac of rat has different characteristics of aspirin with gum compared with that of the control in respect to relative reabsorption of L- histidine.

Everted Intestinal Sacs As In vitro Model For Assessing Absorptivity Of L- Histidine Under The Effect Of Aspirine And Gum Acacia In Male Rats

The purpose of this study was to characterize intestinal permeability changes over a range of physiologically relevant intestinal injury. The experiments were performed in 80 rats subdivided into four groups as aspirin (400 mg/kg b.w.), gum Acacia ( 1g./day) and aspirin with gum Acacia groups for 21 days compared with control group. Relative reabsorption of L- Histidine was greater(p<0.001) in the aspirin in 10 min of incubation compared with that of the control rats. In aspirin in combination with gum Acacia, the relative reabsorption were significantly (p<0.001) decrease in 10, 20 and 30 min. of incubation compared with that of the control rats. Moreover, the relative reabsorption of L-histidine was significantly (p<0.01) reduced by the aspirin at 45 min of time of the incubation buffer compared with that of the control. However, gum acacia treatment was increased at 10 min (p<0.01) ,30 min (p<0.01) and 45 min (p<0.001) respectively compared with that of the control rats. Relative reabsorption of L-histidine record a nonsignificant increase of aspirin at 20 min and 30 min of incubation compared with that of the control. Gum and aspirin with gum at 20min and 45min of incubation resulted an increase and decrease in relative reabsorption of L-histidine respectively compared with that of the control. Aspirin and aspirin in combination with gum acacia treatment increased body, intestinal weights and mucosal total protein significantly with percent changes ranged from 8% to 40% compared with that of the control. On the other hand, gum treatment decreased body, intestinal weights and mucosal total protein significantly with percent changes ranged from 8% to 35% compared with that of the control. These results demonstrated that L-histidine is actively taken up by a gum Acacia system in intestinal everted sac mechanism of rat with energy supplied by glucose and Na + in incubation buffer. Moreover, aspirin system had an inhibitory effect on L-histidine uptake in 45 min of incubation , indicating the saturation by L-histidine in first ten minuts of incubation. Also, these results provide evidence the uptake of L-histidine into rat intestine was not reduced at all by the treatment of aspirin. These results suggest that the uptake of L-histidine by intestinal everted sac of rat has different characteristics of aspirin with gum compared with that of the control in respect to relative reabsorption of L- histidine.

Catalytic activities of cytochrome P450 enzymes and UDP-glucuronosyltransferases involved in drug metabolism in rat everted sacs and intestinal microsomes

1. The use of everted sacs of the small intestine as an enzyme source for studying the metabolism of xenobiotics by cytochrome P450 (P450, CYP) enzymes and UDP-glucuronosyltransferases has been investigated. 2. Most of the drug oxidation activities for testosterone, bufuralol, ethoxyresorufin and 7-ethoxycoumarin resided in the upper part of the everted sacs and in intestinal microsomes. Testosterone 6 β -hydroxylase activities in the everted sacs were about two times higher than those in the intestinal microsomes. By freezing and thawing treatment, the testosterone 6 β - and 16 α -hydroxylase activities of the everted sacs were considerably decreased, and those of the intestinal microsomes were abolished. 3. Microsomal testosterone 6 β -hydroxylation, bufuralol 1'-hydroxylation, and pentoxyresorufin and ethoxyresorufin O -dealkylation were inhibited by ketoconazole, quinine, metyrapone and α -naphthoflavone respectively. Immunoreactive proteins using anti-CYP2B1 and anti-CYP3A2 antibodies were detected in the upper and middle parts of the rat small intestine. 4. Except for morphine 3-glucuronidation, glucuronidation activities in intestinal microsomes or everted sacs were not dependent on the intestinal region. The lower part of the everted sacs exhibited about 10 times higher morphine-3-glucuronidation activities compared with those of the upper part. The glucuronidation activities of 4-nitrophenol in the everted sacs were 10 times higher than those in microsomes. 5. These results demonstrated that the upper part of rat small intestine serves as the major site for intestinal P450-mediated first-pass metabolism. CYP3A enzymes in rat intestinal microsomes may not be stable but probably play an important role in drug oxidations. The high activity of glucuronidation in the rat small intestine should also be considered in terms of drug metabolism.

Hydrolysis by lactase phlorizin hydrolase is the first step in the uptake of daidzein glucosides by rat small intestine in vitro

1. Isoflavones are naturally occurring oestrogenic compounds found in plants, where they exist in the glycosylated form. A proportion of ingested glycosides appears to be absorbed in the upper gastrointestinal tract, where enterocytes play an important role in their metabolism. 2. One hypothesis is that ingestion may involve hydrolysis by the luminally exposed enzyme lactase phlorizin hydrolase (LPH), an enzyme expressed specifically at the small intestinal brush border. 3. Using an everted sac preparation of rat jejunum and an inhibitor of LPH, we investigated the absorption of daidzein- O 7 -glucoside (daidzin) and the effect of LPH inhibition on this process. It was demonstrated that LPH plays a major role in the deglycosylation of daidzin. 4. The hydrolysis product, daidzein, is absorbed by epithelial cells and glucuronidated to daidzein- O 7 -glucuronide, which is subsequently exported primarily to the serosal (vascular) side of the tissue rather than to the luminal side. 5. A small but significant proportion of the intact glycoside is also transferred to the serosal compartment, and in the presence of an LPH inhibitor this was enhanced with a corresponding reduction in deglucosylation and glucuronidation. 6. The results indicate that that LPH plays an important role in the metabolism of glycosylated phytochemicals, and that the expression and activity of this enzyme in the small intestine can modify the profile of metabolites appearing in the circulation.

Tissue-specific regulation of expression and activity of P-glycoprotein in adjuvant arthritis rats

Cyclosporine A and steroids are effective against rheumatoid arthritis and also known as substrates of P-glycoprotein (P-gp). We investigated the effect of arthritis on the hepatic and intestinal P-gp activity in rats, and substantiated the expression level of the hepatic P-gp. Doxorubicin was used as a P-gp substrate. Cumulative biliary excretion and intestinal exsorption of doxorubicin following intravenous administration were compared between adjuvant arthritis (AA) and normal rats. Intestinal P-gp activity was also investigated by intestinal everted sac method, and hepatic P-gp was detected by FITC-labeled antibody and visualized using a confocal laser microscope system. Biliary clearance of doxorubicin in AA rats was significantly decreased from that in normal rats. The expression level of the hepatic P-gp in AA rats was very low compared to normal rats, indicating down-regulation. Intestinal exsorption clearance was not different between AA and normal rats. Permeability of doxorubicin across intestinal everted sac was comparable between AA and normal rats, corresponding to in vivo study. In AA rats, hepatic P-gp activity was decreased due to the reduction of expression level, but intestinal P-gp activity was not changed. Different regulation systems may be involved in liver and intestine.

Roles of the Jejunum and Ileum in the First-Pass Effect as Absorptive Barriers for Orally Administered Tacrolimus

Background. The immunosuppressant tacrolimus shows poor and variable bioavailability following oral administration in clinical use. Recently, the hepatic and intestinal metabolisms, or first-pass effect, of tacrolimus have been suggested to be responsible for its bioavailability. In the present study, we investigated the respective contribution of the jejunum and ileum to the first-pass effect of tacrolimus in rats.Methods. The metabolism of tacrolimus in everted sacs of the duodenum, jejunum, and ileum was examined. Tacrolimus was administered intravenously or intraintestinally to sham-operated, jejunum-resected, or ileum-resected rats. Blood samples were collected over a 240-min period, and whole-blood tacrolimus concentrations were measured by semiautomated microparticle enzyme immunoassay. The pharmacokinetic parameters of tacrolimus in each group were estimated.Results. The metabolic activity of tacrolimus appeared to be the highest in the everted sacs of the duodenum. The bioavailability of tacrolimus in the jejunum- or ileum-resected rats was higher than that in sham-operated controls. On the other hand, the time to peak concentration in the jejunum-resected rats was about twofold slower than those in ileum-resected and sham-operated rats.Conclusions. These results suggested that the first-pass effect of tacrolimus in the small intestine shows regional differences and the extraction of tacrolimus in the small intestine consists of the amount of extraction in the jejunum and ileum. In addition, the ileum rather than the jejunum as a graft of segmental small bowel transplantation would be useful to avoid the adverse effects of tacrolimus.

Adsorption of Cationized Bovine Serum Albumin onto Epithelial Crypt Fractions of the Rat Colon

The purpose of the study was to characterize mucosal attachment of a cationized model protein, bovine serum albumin (BSA), onto the various fractions of colonic crypts epithelium in the rat. BSA was labeled with fluorescein isothiocyanate (FITC) and its surface net electric charge was modified from negative to positive. Attachment of the cationized protein (CF-BSA) onto rat colonic epithelium was performed by incubation of colonic everted sacs in medium containing cationized or non-cationized FITC-labeled BSA. Using a nonenzymatic isolation procedure, colonocytes were harvested from five horizontal fractions of the colonic crypts. BSA adhesion to the isolated colonocytes was quantified spectrofluorometrically. In addition, the effect of increasing concentrations of Mg(2+) on the adsorption of the cationized BSA onto the surface of colonic epithelium was evaluated by measuring its ability to displace the adhered BSA from its binding sites. BSA cationization facilitated protein adherence to the colon epithelium in a crypt depth-dependent manner. The largest extent of adherence was observed in the outer layer (first fraction) of the colon. Binding persisted to approximately half the depth of the crypts. The relation between CF-BSA concentration in the incubation medium and the amount of CF-BSA adsorbed onto the colonic epithelium was exponential in nature. The addition of electrolyte (Mg(2+)) caused a detachment of the CF-BSA. The adsorption process was characterized by Langmuir's adsorption isotherm. It is concluded that cationized BSA could be useful as a targetable drug platform in cases where the target site is the gastrointestinal epithelium.

Ingestion of the soluble dietary fibre, polydextrose, increases calcium absorption and bone mineralization in normal and total-gastrectomized rats

We previously demonstrated that feeding a highly fermentable and water-soluble dietary fibre, guar-gum hydrolysate (GGH) increased intestinal absorption of insoluble Ca salts in total-gastrectomized rats. In the present study, we examined the effects of feeding a less fermentable and water-soluble fibre, polydextrose (PD), on Ca absorption and bone mineralization in the normal and total-gastrectomized rats in comparison with the effects of GGH. Apparent Ca absorption was severely lowered by gastrectomy, and PD feeding (50 g/kg diet) partially restored the reduction of Ca absorption similarly to GGH feeding (50 g/kg diet). PD feeding also increased the Ca absorption in normal rats, but not GGH feeding. Femur Ca concentration was reduced with gastrectomy. Feeding PD for 21 d increased the bone Ca concentration in both normal and gastrectomized rats, but GGH feeding did not. In rats fed PD, pH of the caecal contents was lower than in rats fed fibre-free and GGH diets; however, soluble Ca concentration in the caecal contents was not different between the diet groups. Short-chain fatty acid concentrations were much lower in the PD groups than in the GGH groups. We also examined in vitro Ca absorption by using everted sacs of the small intestine. Addition of PD to the serosal medium of the ileal sacs increased Ca absorption, but addition of GGH did not. These results suggest that the small intestine rather than the large intestine is responsible for the increase in Ca absorption in rats fed PD, and suggests that the mechanism for the increase by PD may be different from that by GGH.

Effects of bioactive substances in milk on mineral and trace element metabolism with special reference to casein phosphopeptides.

Bioactivity of phosphopeptides yielded after tryptic hydrolysis of casein (CPP) was reported more than 50 years ago when CPP were found to improve calcium balance in rachitic newborns. Several investigations have been carried out to study the effects of CPP mainly on calcium metabolism but also on other minerals like iron and zinc. Most of the experiments were in vitro studies or short-term experiments like the effects of CPP after single meals or their effect on mineral disappearance from intestinal everted sac or ligated loop. Investigations on calcium balance were also mainly short term, i.e. 3-4 weeks, and mainly done in rats. A few experiments have been carried out in minipigs, an animal model that is closer to the human than the rat. Studies in human were rare and short term. To date a variety of other peptides have been isolated after enzymatic hydrolysis, and some have been investigated for bioactivity, with equivocal findings. Bioactivity of phosphopeptides seemed to be more obvious when investigations were done in vitro or short term. Results were less clear in metabolic balance studies, especially under physiological conditions. The composition of the basal diet, i.e. content of calcium and phytate, or the protein source had a significant impact on the effect of phosphopeptides. It was concluded that phosphopeptides revealed positive effects on mineral solubility and absorbability, and bone mineralisation under certain experimental conditions. Accordingly they could have a beneficial effect on bone health for some groups of the population.

Use of everted sacs of mouse small intestine as enzyme sources for the study of drug oxidation activities in vitro

1. The use of everted sacs of the small intestine as an enzyme source for the study of the first-pass metabolism of xenobiotics by cytochrome P450s (P450, CYP) is described. Several drug oxidation activities for testosterone, chlorzoxazone, tolbutamide, bufuralol and warfarin were observed when everted sacs (1-cm segment) from different parts ofmouse small intestine were incubated with an NADPH-generating system and each substrate. 2. Most of the drug hydroxylase activities resided in the upper part of mouse small intestine and these activities were much higher than those of intestinal microsomes. Drug oxidation activities decreased along the distance from the upper part of the small intestine except for warfarin hydroxylation. 3. Testosterone 6β-hydroxylation in the everted sacs exhibited the highest catalytic activities among the drug oxidations tested here. In the upper part of the small intestine, thetestosterone 6β-hydroxylase activities of everted sacs subjected once to freezing and thawing were substantially decreased compared with the untreated everted sacs. 4. Testosterone 6β-hydroxylase activities in the everted sacs of the small intestine were significantly inhibited by ketoconazole. Immunoreactive proteins using anti-CYP3A antibodies were detected in the upper and middle parts of the small intestine. 5. The results demonstrated that the upper part of the mouse small intestine serves as the major site for intestinal P450 mediated first-pass metabolism. Everted sacs of the small intestine are therefore useful for the study of drug metabolism as well as of transport and absorption.

Effect of portal hypertension caused by chronic high venous pressure on small-intestinal sugar absorption

The effect of portal hypertension and chronic high venous pressure on the absorption of the small intestine was examined by constricting the suprahepatic and subdiaphragmatic inferior vena cava in rats. A group of rats with the constricted suprahepatic and subdiaphragmatic inferior vena cava comprised group 1 ( n = 9) and another group of rats with only laparotomy comprised group 2 ( n = 9). Two months after the operation, sugar absorption and other parameters were measured. The blood pressures of the infrahepatic inferior vena cava and the portal vein 8 wk after the operation in group 1 were significantly higher than those in group 2. The results of d-xylose absorption tests showed that the amount of excreted d-xylose in urine in group 1 was significantly lower than that of group 2, but the d-xylose everted sac test showed no significant differences between the two groups. The glucose everted sac test showed that the amount of glucose absorption in group 1 was significantly lower than that in group 2. These findings suggest that chronic high venous pressure caused by constriction of suprahepatic inferior vena cava may lead to sugar malabsorption. We conclude that portal hypertension with high venous pressure below the diaphragm can lead to sugar malabsorption in the intestine.

Effects of Magnesium on Calcium Absorption in the Small Intestine of Goat and Sheep in vitro

In the first experiment, the effect of mucosal high Mg concentration on Ca absorption in everted sacs of the duodenum, jejunum and ileum of goats and sheep was examined. An increase in mucosal MgCl2 concentration from 1.2 to 60.0mmol/l significantly increased Ca absorption (P<0.05) in the duodenum and ileum of goats and in the jejunum and ileum of sheep. In the second experiment, it was observed that Mg-induced Ca absorption was linearly dependent on mucosal Mg concentrations (r=0.744, P<0.001), and it was the mucosal and not the serosal solution that significantly increased Ca absorption (P<0.05) in the ovine ileum. In addition, potential difference (PD) between mucosal and serosal sides did not correlate with Mg-induced Ca absorption. In the third experiment, it was found that mucosal 60.0mmol/l MgCl2 significantly increased Jms (unidirectional flux rate from mucosal to serosal)of45 Ca2+ (P<0.05) in the caprine ileum. From these findings, it is concluded that the ileum of goats and sheep is a preferable site to observe the Mg-induced Ca absorption in vitro, and that Mg exerts its action on a mucosal and not a serosal side of the epithelial tissue without depending on PD.

Transition metal saccharide chemistry and biology: syntheses, characterization, solution stability and putative bio-relevant studies of iron–saccharide complexes

A number of Fe(III) complexes of saccharides and their derivatives, and those of ascorbic acid were synthesized, and characterized by a variety of analytical, spectral (FT-IR, UV–Vis, EPR, Mössbauer and EXAFS), magnetic and electrochemical techniques. Results obtained from various methods have shown good correlations. Data obtained from EPR, magnetic susceptibility and EXAFS techniques could be fitted well with the mono-, di- and trinuclear nature of the complexes. The solution stability of these complexes has been established using UV–Vis absorption and cyclic voltammetric techniques as a function of pH of the solution. Mixed valent, Fe(II,III) ascorbate complexes have also been synthesized and characterized. Reductive release of Fe(II) from the complexes using sodium dithionite has been addressed. In vitro absorption of Fe(III)–glucose complex has been studied using everted sacs of rat intestines and the results have been compared with that of simple ferric chloride. Fe(III)–saccharide complexes have shown regular protein synthesis even in hemin-deficient rabbit reticulocyte lysate indicating that these complexes play a role that is equivalent to that played by hemin in order to restore the normal synthesis of protein. These complexes have exhibited enhanced DNA cleavage properties in the presence of hydrogen peroxide with pUC-18 DNA plasmid.

Correlation of two bioadhesion assays: the everted sac technique and the CAHN microbalance

This contribution correlates two in vitro methods utilized to determine bioadhesion. One method, the everted intestinal sac technique, is a passive test for bioadhesion involving several polymer microspheres and a section of everted intestinal tissue. The other method, the CAHN microbalance, employs a CAHN dynamic contact angle analyzer with modified software to record the tensile forces measured as a single polymer microsphere is pulled from intestinal tissue. This study demonstrates that CAHN and everted sac experiments yield similar results when used to quantify the bioadhesive nature of polymer microsphere systems. A polymer showing high adhesion in one method also demonstrates high bioadhesion in the other method; polymers that exhibit high fracture strength and tensile work measurements with the CAHN microbalance also yield high binding percentages with the everted sac method. The polymers tested and reported here are poly(caprolactone) and different copolymer ratios of poly(fumaric-co-sebacic anhydride). The results of this correlation demonstrate that each method alone is a valuable indicator of bioadhesion.

Gastrointestinal absorption of drugs: methods and studies.

Physico-chemical descriptors of drug molecules are often not adequate in predicting their oral bioavailability. In vitro methods can be useful in evaluating some of the different factors contributing to bioavailability. While physical parameters such as drug solubility may effect oral bioavailability, in most cases, the major determining factors are likely to be metabolism, and absorption at the intestinal level. Metabolism may be preabsorptive, as with peptides, or during absorption, particularly as a result of the activity of the intracellular enzyme CYP3A4. Absorption may be transcellular (membrane diffusion, carrier-mediated, endocytosis) or paracellular, while p-glycoprotein activity in the apical cell membrane may limit bioavailability by expelling drugs from the mucosal cells. Knowledge of the absorption mechanism is important in determining formulation strategies. The different in vitro techniques used to study absorption have advantages and disadvantages. Ussing chambers can be useful to measure bidirectional transport, but most studies use simple salt media, and full tissue viability is doubtful. Caco-2 cell monolayers are human cells, but the system is static, and gives very low rates of transport, and exagerated enhancement of the paracellular route compared with small intestine. The rat everted gut sac incubated in tissue culture medium maintains tissue viability and gives reliable data, although it is a closed system. In situ perfusion gives no information on events at the cellular level, and absorption may be reduced by anaesthesia and surgical manipulation. In vivo perfusion in man, with multichannel tubes, gives valuable data, but is not practical for screening. Pharmacokinetic modelling can also give useful data such as the existence of different absorption sites. Permeability values from the literature show that for small hydrophilic molecules, which pass by the paracellular route, the improved everted sac gives values close to those for humans, while values with Caco-2 cells are orders of magnitude lower.