Differential Recognition of ACE Inhibitors in Xenopus Laevis Oocytes Expressing Rat PEPT1 and PEPT2

Purpose. To examine the mechanism of inhibition of glycylsarcosine(GlySar) transport by quinapril and enalapril, and whether or notangiotensin converting enzyme (ACE) inhibitors are transported by PEPT2as well as by PEPT1. Methods. Xenopus laevis oocytes were cRNA-injected with rat PEPT1or PEPT2 and the transport kinetics of radiolabeled GlySar were studiedin the absence and presence of quinapril and enalapril. Thetwo-microelectrode voltage-clamp technique was also performed to probe theelectrogenic uptake of captopril, quinapril and enalapril. Results. Kinetic analyses demonstrated that quinapril inhibited theuptake of GlySar in a noncompetitive manner in Xenopus oocytesinjected with PEPT1 or PEPT2 (Ki = 0.8 or 0.4 mM, respectively).In contrast, a competitive interaction was observed between GlySarand enalapril (Ki = 10.8 mM for PEPT1 or 4.3 mM for PEPT2).Most significantly, captopril and enalapril, but not quinapril, inducedinwardly-directed currents in both PEPT1- and PEPT2-expressedoocytes. Conclusions. These results are unique in providing direct evidence forthe substrate recognition and transport of some ACE inhibitors by thehigh- and low-affinity oligopeptide transporters. Our findings point todifferences between PEPT1 and PEPT2 in their affinity to, rather thanin their specificity for, ACE inhibitors.     

PEPT2-Mediated Uptake of Neuropeptides in Rat Choroid Plexus

Purpose. The peptide transporter PEPT2 was recently shown to be functionally active in rat choroid plexus, suggesting that it may play a role in neuropeptide homeostasis in the cerebrospinal fluid. This study, therefore, examined the role of PEPT2 in mediating neuropeptide uptake into choroid plexus. Methods. Whole-tissue rat choroid plexus uptake studies were performed on GlySar in the absence and presence of neuropeptides and on carnosine. Results. The neuropeptides NAAG, CysGly, GlyGln, kyotorphin, and carnosine inhibited the uptake of radiolabeled GlySar at 1.0 mM concentrations. In contrast, TRH, [D-Arg²]-kyotorphin, glutathione, and homocarnosine did not inhibit GlySar uptake. Kyotorphin, an analgesic, was a competitive inhibitor of GlySar with a Ki of 8.0 M. The direct uptake of carnosine was also shown to be mediated by PEPT2 in isolated choroid plexus (Km = 39.3 M; Vmax = 73.9 pmol/mg/min). Radiolabeled carnosine uptake was inhibited by 1.0 mM concentrations of GlySar or carnosine but not homocarnosine, L-histidine, or -alanine. Conclusions. These findings indicate that PEPT2 mediates the uptake of a diverse group of neuropeptides in choroid plexus, and suggests a role for PEPT2 in the regulation of neuropeptides, peptide fragments, and peptidomimetics in cerebrospinal fluid.     

Role of PEPT2 in the Choroid Plexus Uptake of Glycylsarcosine and 5-Aminolevulinic Acid: Studies in Wild-Type and Null Mice

PURPOSE: To determine the importance of PEPT2 in the uptake of glycylsarcosine (GlySar) and 5-aminolevulinic acid (5-ALA) in mouse choroid plexus whole tissue. METHODS: Uptake studies were performed in bicarbonate artificial cerebrospinal fluid buffer using choroid plexuses isolated from PEPT2+/+ and PEPT2-/- mice. [14C]GlySar and [14C]5-ALA were studied as a function of temperature, concentration, potential inhibitors, and low sodium conditions. RESULTS: PEPT2-/- mice exhibited a 90% reduction in GlySar uptake (p < 0.001) and a 92% reduction in 5-ALA uptake (p < 0.001) as compared to wild type animals. At 4 degrees C (vs. 37 degrees C), GlySar uptake was reduced by 95% in PEPT2+/+ mice; no difference was observed in null animals. Unlabeled GlySar inhibited the uptake of [14C]GlySar in PEPT2+/+ mice (p < 0.01); self-inhibition did not occur in PEPT2-/- mice. GlySar demonstrated saturable uptake in PEPT2+/+ mice (Vmax = 16.4 pmol mg(-1) min(-1), Km = 70 microM, Kd = 0.014 microl mg(-1) min(-1)), however, uptake was linear in PEPT2-/- mice (Kd = 0.023 microl mg(-1) min(-1)). Low sodium buffer (1 mM) resulted in 75% and 59% reductions, respectively, in GlySar (p < 0.001) and 5-ALA (p < 0.01) uptake in PEPT2+/+ mice; no differences were observed in PEPT2-/- mice. Overall, about 90-95% of the choroid plexus uptake of GlySar and 5-ALA was mediated by PEPT2, with about 5-10% of the residual uptake occurring by nonspecific mechanisms. CONCLUSIONS: The results demonstrate that PEPT2 is the only transporter responsible for the choroid plexus uptake of GlySar and 5-ALA. They also suggest a role for PEPT2 in the clearance of dipeptides and endogenous peptidomimetics from cerebrospinal fluid.     

Floxuridine Amino Acid Ester Prodrugs: Enhancing Caco-2 Permeability and Resistance to Glycosidic Bond Metabolism

Purpose The aim of this study was to synthesize amino acid ester prodrugs of 5-fluoro-2′-deoxyuridine (floxuridine) to enhance intestinal absorption and resistance to glycosidic bond metabolism.Methods Amino acid ester prodrugs were synthesized and examined for their hydrolytic stability in human plasma, in Caco-2 cell homogenates, and in the presence of thymidine phosphorylase. Glycyl-l-sarcosine uptake inhibition and direct uptake studies with HeLa/PEPT1 cells [HeLa cells overexpressing oligopeptide transporter (PEPT1)] were conducted to determine PEPT1-mediated transport and compared with permeability of the prodrugs across Caco-2 monolayers.Results Isoleucyl prodrugs exhibited the highest chemical and enzymatic stability. The prodrugs enhanced the stability of the glycosidic bond of floxuridine. Thymidine phosphorylase rapidly cleaved floxuridine to 5-fluorouracil, whereas with the prodrugs no detectable glycosidic bond cleavage was observed. The 5′-l-isoleucyl and 5′-l-valyl monoester prodrugs exhibited 8- and 19-fold PEPT1-mediated uptake enhancement in HeLa/PEPT1 cells, respectively. Uptake enhancement in HeLa/PEPT1 cells correlated highly with Caco-2 permeability for all prodrugs tested. Caco-2 permeability of 5′-l-isoleucyl and 5′-l-valyl prodrugs was 8- to 11-fold greater compared with floxuridine.Conclusions Amino acid ester prodrugs such as isoleucyl floxuridine that exhibit enhanced Caco-2 transport and slower rate of enzymatic activation to parent, and that are highly resistant to metabolism by thymidine phosphorylase may improve oral delivery and therapeutic index of floxuridine.     

Peptide Transporter 1 Is Responsible for Intestinal Uptake of the Dipeptide Glycylsarcosine: Studies in Everted Jejunal Rings from Wild-type and Pept1 Null Mice

The purpose of this study was to determine the relative importance of peptide transporter 1 (PEPT1) in the uptake of peptides/mimetics from mouse small intestine, using glycylsarcosine (GlySar). After isolating jejunal tissue from wild-type and Pept1 null mice, 2 cm intestinal segments were everted and mounted on glass rods for tissue uptake studies. [¹⁴C]GlySar (4 μM) was studied as a function of time, temperature, sodium and pH, concentration, and potential inhibitors. Compared with wild-type animals, Pept1 null mice exhibited a 78% reduction in GlySar uptake at pH 6.0 at 37°C. GlySar uptake showed pH dependence, with peak values between pH 6.0 and 6.5 in wild-type animals, whereas no such tendency was observed in Pept1 null mice. GlySar exhibited Michaelis–Menten uptake kinetics and a minor nonsaturable component in wild-type animals. In contrast, GlySar uptake occurred only by a nonsaturable process in Pept1 null mice. GlySar uptake was significantly inhibited by dipeptides, aminocephalosporins, angiotensin-converting enzyme inhibitors, and the antiviral prodrug valacyclovir; these inhibitors had little, if any, effect on the uptake of GlySar in Pept1 null mice. The findings demonstrate that PEPT1 plays a critical role in the uptake of GlySar in jejunum and suggest that PEPT1 is the major transporter responsible for the intestinal absorption of small peptides.     

Expression of Multidrug Resistance-Associated Protein (MRP) in Human Retinal Pigment Epithelial Cells and Its Interaction with BAPSG,a Novel Aldose Reductase Inhibitor

Purpose. The objective of this study was to determine the expression and activity of multidrug resistance–associated protein (MRP) in the retinal pigment epithelial (RPE) cells and to further assess whether BAPSG, a novel anionic aldose reductase inhibitor, interacts with MRP. Methods. Functional and biochemical evidence for MRP was obtained in a human retinal pigment epithelial (ARPE–19) cell line and primary cultures of human retinal pigment epithelial (HRPE) cells. Fluorescein accumulation and efflux in the presence and absence of MRP inhibitors was used to obtain functional evidence for MRP. Western blots and RT–PCR were used to obtain biochemical evidence for MRP1. The influence of MRP inhibitors on BAPSG accumulation and efflux in ARPE–19 cells was determined to understand its interaction with MRP. Results. MRP inhibitors increased fluorescein accumulation and reduced efflux in RPE cells. Both cell types exhibited a 190–kDa western blot band corresponding to MRP1 protein and a 287 bp RT–PCR band corresponding to MRP1 mRNA. MRP inhibitors reduced BAPSG efflux and increased its accumulation in ARPE–19 cells. Conclusions. MRP is functionally and biochemically active in human RPE cells. Anionic BAPSG is a likely substrate for MRP.     

Protective effects of melatonin and memantine in human retinal pigment epithelium (ARPE-19) cells against 2-ethylpyridine-induced oxidative stress: implications for age-related macular degeneration

Purpose: To investigate the possible protective effects of melatonin and memantine (MMT) against 2-ethylpyridine (2-EP)-induced oxidative stress and mitochondrial dysfunction in human RPE (ARPE-19) cells in vitro.

Materials and methods: The ARPE-19 cells were divided into seven groups. Oxidative stress was triggered by incubating the ARPE-19 cells with 30?μM of 2-EP for 24?h. Then, 200?μM of melatonin was administered over three days and 20?μM of MMT over six hours prior to the experiment. The effects of melatonin and MMT on the intracellular calcium release mechanism, reactive oxygen species production, caspase-3 and caspase-9 activities, as well as vascular endothelial growth factor levels were measured.

Results: Melatonin and MMT were found to significantly decrease apoptosis levels. The intracellular calcium release was regulated by both melatonin and MMT. Further, melatonin and MMT significantly decreased both caspase-3 and caspase-9 activities, as well as pro-caspase and poly(ADP-ribose) polymerase expression, in ARPE-19 cells. Moreover, melatonin significantly increased the protective effect of MMT. The combination of melatonin and MMT significantly decreased 2-EP-induced oxidative toxicity and apoptosis by inhibiting the intracellular reactive oxygen species production and mitochondrial depolarization levels.

Conclusions: These notable findings are the first to demonstrate the synergistic protective effects of melatonin and MMT against 2-EP-induced oxidative stress in ARPE-19 cells.     

Lutein improves cell viability and reduces Alu RNA accumulation in hydrogen peroxide challenged retinal pigment epithelial cells

Purpose: Dysfunction of the microRNA (miRNA)-processing enzyme DICER1 and Alu RNA accumulation are linked to the pathogenesis of age-related macular degeneration (AMD). This study determined the optimal dose of lutein (LUT) and zeaxanthin (ZEA) to protect human retinal pigment epithelium (RPE) cells against hydrogen peroxide (H₂O₂). The effect of the optimal dose of LUT and ZEA as DICER1 and Alu RNA modulators in cultured human RPE cells challenged with H₂O₂ was investigated.

Materials and methods: ARPE-19 cells were pre-treated with LUT, ZEA, or both for 24?h before 200?μM H₂O₂ challenge. Cell viability was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. DICER1 and Alu RNA were quantified by western blotting and real-time polymerase chain reaction, respectively.

Results: H₂O₂ increased cell Alu RNA expression and decreased cell viability of ARPE-19, but had no significant impact on the DICER1 protein level. LUT, alone and in combination with ZEA pre-treatment, prior to H₂O₂ challenge significantly improved cell viability of ARPE-19 and reduced the level of Alu RNA compared to the negative control.

Conclusions: These results support the use of LUT alone, and in combination with ZEA, in AMD prevention and treatment. This study is also the first to report LUT modulating effects on Alu RNA.     

在体单向肠灌流模型研究帕拉米韦拟肽类衍生物的大鼠小肠吸收特性

目的研究帕拉米韦及其拟肽类衍生物的大鼠小肠吸收机制,筛选出膜渗透性最大的衍生物。方法采用大鼠在体单向灌流法研究帕拉米韦拟肽类衍生物的小肠吸收,采用高效液相色谱法测定药物和酚红的浓度。建立lgD预测值和lgP之间的关系。结果帕拉米韦拟肽类衍生物的膜渗透系数都比帕拉米韦高,其中帕拉米韦L-异亮氨酸衍生物具有最高的膜渗透性;寡肽转运蛋白(PEPT1)典型底物甘氨酰肌氨酸能显著降低帕拉米韦拟肽类衍生物的小肠吸收,而L-缬氨酸不具有这种能力。结论帕拉米韦拟肽类衍生物是PEPT1的底物,它们在大鼠小肠内的吸收是PEPT1介导的主动转运过程。     

Species-dependent uptake of glycylsarcosine but not oseltamivir in Pichia pastoris expressing the rat, mouse, and human intestinal peptide transporter PEPT1

The purpose of this study was to determine whether glycylsarcosine (a model dipeptide) and oseltamivir (an antiviral prodrug) exhibited a species-dependent uptake in yeast Pichia pastoris expressing the rat, mouse, and human homologs of PEPT1. Experiments were performed with [(3)H]glycylsarcosine (GlySar) in yeast P. pastoris expressing human, mouse, and rat peptide transporter 1 (PEPT1), in which uptake was examined as a function of time, concentration, potential inhibitors, and the dose-response inhibition of GlySar by oseltamivir. Studies with [(14)C]oseltamivir were also performed under identical experimental conditions. We found that GlySar exhibited saturable uptake in all three species, with K(m) values for human (0.86 mM) > mouse (0.30 mM) > rat (0.16 mM). GlySar uptake in the yeast transformants was specific for peptides (glycylproline) and peptide-like drugs (cefadroxil, cephradine, and valacyclovir), but was unaffected by glycine, l-histidine, cefazolin, cephalothin, cephapirin, acyclovir, 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid, tetraethylammonium, and elacridar. Although oseltamivir caused a dose-dependent inhibition of GlySar uptake [IC(50) values for human (27.4 mM) > rat (18.3 mM) > mouse (10.7 mM)], the clinical relevance of this interaction would be very low in humans. Of importance, oseltamivir was not a substrate for the intestinal PEPT1 transporter in yeast expressing the three mammalian species tested. Instead, the prodrug exhibited nonspecific binding to the yeast vector and PEPT1 transformants. Finally, the mouse appeared to be a better animal model than the rat for exploring the intestinal absorption and pharmacokinetics of peptides and peptide-like drugs in human.     

Intestinal Transport of β-Lactam Antibiotics: Analysis of the Affinity at the H+/Peptide Symporter (PEPT1), the Uptake into Caco-2 Cell Monolayers and the Transepithelial Flux

Purpose. This study on the intestinal transport of -lactam antibiotics was undertaken to investigate the correlation between cellular transport parameters and the bioavailability. Methods. Transport of 23 -lactam antibiotics was characterized by measuring their ability to inhibit the uptake of glycylsarcosine into Caco-2 cells, their uptake into the cells and their total flux across the cell monolayers. Results. Ceftibuten and cyclacillin were recognized by PEPT1 with affinity constants comparable to those of natural dipeptides (K_i = 0.3 and 0.5 mM, respectively). Cefadroxil, cefamandole, cephradine, cefaclor, cefuroxime-axetil, cefixime, cephalotin, cephalexin and ampicillin also interacted with PEPT1 (K_i = 7-14 mM). In contrast, cefapirin, cefodizime, cefuroxime, cefmetazole, ceftazidime, benzyl-penicillin, ceftriaxone, cefpirome, cefotaxime, cefepime, cephaloridine and cefsulodin displayed no affinity to the transport system (K_i > 20 mM). The uptake into the cells and the transepithelial flux was highest for those -lactam antibiotics, which showed the strongest inhibition of [¹⁴C]Gly-Sar transport (p < 0.0001). Exceptions were cefuroxim-axetil and cephalotin. Conclusions. The probability of oral bioavailability for -lactam antibiotics is mainly determined by their affinity to PEPT1. A threshold K_i value of 14 mM with respect to Gly-Sar uptake is required.     

5′-Amino Acid Esters of Antiviral Nucleosides, Acyclovir, and AZT Are Absorbed by the Intestinal PEPT1 Peptide Transporter

Purpose. General use of nucleoside analogues in the treatment of viral infections and cancer is often limited by poor oral absorption. Valacyclovir, a water soluble amino acid ester prodrug of acyclovir has been reported to increase the oral bioavailability of acyclovir but its absorption mechanism is unknown. This study characterized the intestinal absorption mechanism of 5-amino acid ester prodrugs of the antiviral drugs and examined the potential of amino acid esters as an effective strategy for improving oral drug absorption. Methods. Acyclovir (ACV) and Zidovudine (AZT) were selected as the different sugar-modified nucleo-side antiviral agents and synthesized to L-valyl esters of ACV and AZT (L-Val-ACV and L-Val-AZT), D-valyl ester of ACV (D-Val-ACV) and glycyl ester of ACV (Gly-ACV). The intestinal absorption mechanism of these 5-amino acid ester prodrugs was characterized in three different experimental systems; in siturat perfusion model, CHO/hPEPTl cells and Caco-2 cells. Results. Testing 5-amino acid ester prodrugs of acyclovir and AZT, we found that the prodrugs increased the intestinal permeability of the parent nucleoside analogue 3- to 10-fold. The dose- dependent permeation enhancement was selective for the L-amino acid esters. Competitive inhibition studies in rats and in CHO cells transfected with the human peptide transporter, hPEPTl, demonstrated that membrane transport of the prodrugs was mediated predominantly by the PEPT1 H⁺/dipeptide cotransporter even though these prodrugs did not possess a peptide bond. Finally, transport studies in Caco-2 cells confirmed that the 5-amino acid ester prodrugs enhanced the transcellular transport of the parent drug. Conclusions. This study demonstrates that L-amino acid-nucleoside chimeras can serve as prodrugs to enhance intestinal absorption via the PEPT1 transporter, providing a novel strategy for improving oral therapy of nucleoside drugs.     

Human Dipeptide Transporter,hPEPTl, Stably Transfected into Chinese Hamster Ovary Cells

Purpose. A cDNA encoding the H⁺-coupled peptide transporter, hPEPTl, has previously been cloned from human ileum (8). The objective of this study was to establish a stably transfected cell line expressing hPEPTl in mammalian cell culture. Methods. The hPEPTl cDNA was subcloned into an expression vector carrying the CMV promoter and a neomycin resistance gene. This vector, pCDNA3-PEPT1, was transiently transfected into several cell lines to identify those capable of expressing PEPT1 transport function. CHO cells were selected and stably transfected with PEPT1 (CHO-PEPT1). Dipeptide transport activity was measured with ³H-Gly-Sar, in the presence and absence of inhibitors. Results. The clonal cell line, CHO-PEPT1, displayed high transport activity. Dipeptide transport was sensitive to pH and specific for dipeptides and other small peptides. Peptidomimetic antibiotics, such as cephalexin, were competitors for peptide transport. Conclusions. The stably transfected cell line, CHO-PEPT1 exhibits enhanced transport over that of cell lines with native expression of PEPT1, and therefore, represents a useful tool for rapid screening of drugs that utilize the peptide transporter in the human intestine for absorption.     

Silk fibroin nanoparticles for enhanced bio-macromolecule delivery to the retina

The aim of this study was to investigate intravitreal injection of silk fibroin nanoparticles (SFNs) encapsulating bio-macromolecules, achieving enhanced drug bioavailability, and extended retention in retina. SFNs were prepared with regenerated silk fibroin using desolvation method with fluorescein isothiocyanate labeled bovine serum albumin (FITC-BSA) as bio-macromolecular model drug encapsulated. In vitro physicochemical properties and in vitro drug release of FITC-BSA loaded SFNs (FITC-BSA-SFNs) were evaluated. Cytotoxicity, cellular uptake, and retention of FITC-BSA-SFNs were determined in human retinal pigment epithelial cell line (ARPE-19). In addition, in vivo distribution and safety of intravitreally administered FITC-BSA-SFNs were investigated in New Zealand white rabbits. The particle size of FITC-BSA-SFNs was 179.1?±?3.7?nm with polydispersity index of 0.102?±?0.033 and the zeta potential was greater than ?25?mV. FITC-BSA-SFNs exhibited excellent biocompatibility with no cytotoxicity observed within 24 and 48?h in AREP-19 cells. Compared to FITC-BSA solution, FITC-BSA-SFNs showed enhanced cellular uptake and prolonged retention. Furthermore, FITC-BSA-SFNs achieved accumulated distribution and extended retention in retina in vivo following intravitreal injection compared to a single administration of free drug solution. Therefore, this bio-macromolecule delivery platform based on SFNs could have great potential in the treatment of posterior segment disorders.     

Inhibitory Effect of Zinc on PEPT1-Mediated Transport of Glycylsarcosine and β-Lactam Antibiotics in Human Intestinal Cell Line Caco-2

Purpose. The aim of this study was to examine the effects of zinc on the intestinal peptide transporters (PEPT1 and basolateral peptide transporter) and to elucidate the mechanism of the interactions. Methods. Caco-2 cells were pretreated with zinc, and the uptake studies were carried out. Results. Zinc treatment resulted in the inhibition of [¹⁴C]glycylsarcosine (Gly-Sar) uptake via PEPT1 in a concentration-dependent manner, whereas it showed moderate inhibitory effect on the basolateral peptide transporter. Zinc also inhibited the uptake of oral -lactam antibiotics such as ceftibuten and cephradine by PEPT1. Kinetic analysis showed that zinc treatment increased K _m values without affecting V _max values of the [¹⁴C]Gly-Sar uptake. The inhibition of [¹⁴C]Gly-Sar uptake induced by zinc was observed in the presence of an H⁺ gradient but not in the absence of an H⁺ gradient. Conclusions. These results indicate that zinc is a competitive inhibitor of PEPT1. Zinc inhibited the PEPT1 function, possibly by interacting with histidine residues of PEPT1 that are part of an H⁺-binding site. These findings would provide important information for clinical, physiologic, and biochemical aspects of peptide transporters.     

Glycyl-<Emphasis Type="SmallCaps">l</Emphasis>-Glutamine Disposition in Rat Choroid Plexus Epithelial Cells in Primary Culture: Role of PEPT2

Purpose The purpose of this research was to determine the polarity and directionality of the PEPT2-mediated uptake and transepithelial transport of the neuropeptide glycyl-l-glutamine (GlyGln) in choroid plexus.Methods The transport kinetics of [³H]GlyGln was studied in neonatal rat choroid plexus epithelial cells in primary culture grown on laminin-coated Transwell filter inserts. Using a bicarbonate artificial cerebrospinal fluid (CSF) buffer (pH 7.4) at 37°C, GlyGln studies were performed as a function of time, substrate concentration, and the presence of potential inhibitors (at 1 mM).Results GlyGln (2 μM) accumulation was about three to four times greater when introduced from the apical (CSF-facing) as opposed to the basal (blood-facing) side of the cell monolayer, and transepithelial transport was about two times greater in the apical-to-basal direction. The apical uptake of radiolabeled GlyGln (2 μM) was inhibited significantly by dipeptides (i.e., unlabeled GlyGln and cysteinylglycine) and some neuropeptides (i.e., carnosine, N-acetylaspartylglutamate, kyotorphin), but was unaffected by amino acids (i.e., glycine, glutamine) as well as by [d-Arg²]-kyotorphin and glutathione. The concentration-dependent apical uptake of GlyGln (2–1000 μM) was characterized by a high-affinity process (i.e., V_max of 72 pmol/mg/min; K_m of 136 μM), consistent with the properties of PEPT2. The intracellular hydrolysis of GlyGln was extensive, however, with only 40% of the dipeptide remaining intact after 1 h.Conclusions The results demonstrate that PEPT2 plays an important role in regulating the apical uptake of GlyGln at the blood–CSF interface. Once inside the cell, GlyGln is rapidly degraded to its constitutive amino acids for further processing.     

Quantitative Evaluation of PEPT1 Contribution to Oral Absorption of Cephalexin in Rats

Purpose  PEPT1 mediates the intestinal absorption of many drugs, but its contribution to oral absorption of drugs is still controversial. The objective of this study is to quantitatively evaluate the contribution of PEPT1 to oral absorption of cephalexin, a typical substrate for PEPT1, in rats. Materials and Methods  The absorbability of cephalexin via PEPT1 or passive diffusion was assessed in five intestinal segments by utilizing glycyl-proline as a competitive inhibitor by in-situ closed loop method. Absorption kinetics of cephalexin after oral administration was predicted by GI-Transit-Absorption model. Results  Absorbability of cephalexin was segment-dependent, and concentration-dependent in all the segments except for the lower ileum. Intrinsic absorption rate constant via PEPT1 ranged from 0.64 to 4.07 h⁻¹. The absorption rate constants via passive diffusion ranged from 0.78 to 1.24 h⁻¹. Plasma concentration–time profile of cephalexin was successfully predicted and the substantial contribution of PEPT1 to the oral absorption was calculated to be from 46% to 60% of total absorption. Simulation study indicated that 83% bioavailability would be expected for cephalexin even though PEPT1 does not function. Conclusions  PEPT1 substantially contributes to oral absorption of cephalexin, around a half of total absorption. However, the function of PEPT1 can be compensated by passive diffusion for cephalexin.