Transport of proteins and peptides across human cultured alveolar A549 cell monolayer

An in vitro cultured monolayer system of alveolar epithelial cells was used as a model to investigate the transport pathway of the peptides and proteins, salmon calcitonin (sCT), insulin (INS), recombinant hirudin (rHAV2), and recombinant human growth hormone (rhGH), in pulmonary epithelium. Human lung adenocarcinoma A549 cells formed continuous monolayers when grown on the polycarbonate filters of Transwell plates. The transport of the peptides and proteins having MW of 3400-22,000 Da was studied under different conditions. The results showed that the apparent permeability coefficients (P(app)) of these macromolecules across A549 cell monolayers ranged from 2x10(-6) to 5x10(-6) cm s(-1) and exhibited a good inverse correlation with molecular weight. No concentration, direction, or temperature dependence was observed in the permeation of sCT, INS, and rHAV2. While the P(app) of rhGH in the BA direction (2.25x10(-6) cm s(-1)) was less than that in the AB direction at both concentrations (3.20x10(-6) and 3.29x10(-6) cm s(-1)). The P(app) values of rhGH were concentration and temperature independent in the AB direction. These findings suggest that the hydrophilic peptides and proteins used in this study, sCT, INS, rHAV2, and rhGH, appear to cross the A549 cell monolayers via a paracellular pathway by a passive diffusion mechanism.     

应用A549细胞单层模型研究蛋白多肽类药物肺部吸收的特性应用A549细胞单层模型研究蛋白多肽类药物肺部吸收的特性

药物通过呼吸进行吸收的路径依次为气管→支气管→肺泡,其中通过肺泡上皮细胞吸收占90％以上。由于肺泡上皮细胞使大分子的蛋白多肽类药物不易通过,是它们渗透的主要屏障,因此对肺泡膜屏障特性、药物透过过程及渗透特性的研究有助于更     

Sulfasalazine transport in in-vitro, ex-vivo and in-vivo absorption models: contribution of efflux carriers and their modulation by co-administration of synthetic nature-identical fruit extracts

Sulfasalazine is characterised by low oral bioavailability. In this study, its intestinal transport characteristics were studied in an in-vitro, ex-vivo and in-situ system. The absorptive transport of sulfasalazine across Caco-2 monolayers appeared to be lower than the secretory transport (P(app-abs) = 0.21 +/- 0.02 x 10(-6) cm s(-1) and P(app-secr) = 2.97 +/- 0.30 x 10(-6) cm s(-1), respectively). This polarity in transport of sulfasalazine was not mediated by P-glycoprotein (P-gp), as inclusion of verapamil (100 microM) did not have any effect on the transport polarity of sulfasalazine. However, inclusion of the multidrug resistance-associated protein (MRP) inhibitors benzbromarone (50 microM) and sulfinpyrazone (1 mM), and the glutathione-depleting agent chlorodinitrobenzene (100 microM), resulted in an increased absorptive transport of sulfasalazine in the Caco-2 system (P(app-abs) = 0.64 +/- 0.02, 0.51 +/- 0.04 and 0.60 +/- 0.03 x 10(-6) cm s(-1), respectively). The interference of carriers implies that, during absorption, interactions with food components may occur at the level of this carrier. Therefore, the effect of food extracts was studied in a parallel set of experiments. For two standardized nature-identical fruit extracts (pineapple and apricot extract) a concentration-dependent absorption-enhancing effect could be observed in the Caco-2 system. The functional expression of similar carriers was also demonstrated in rat ileum in the Ussing chamber system. Interaction studies with fruit extracts in the Ussing chamber system, as well as in the in-situ intestinal perfusion study, revealed a 2- to 4-fold increase in the absorptive transport of sulfasalazine. These results indicate that food components in the intestinal lumen can have a significant impact on the intestinal absorption characteristics of sulfasalazine by modulating the biochemical barrier function of the intestinal mucosa.     

Chlorzoxazone, a selective probe for phenotyping CYP2E1 in humans.

The ability of human cytochromes P450 other than CYP2E1 to catalyse the 6-hydroxylation of chlorzoxazone (6-OH-CHZ) was examined in vitro using human liver microsomal preparations and in vivo using chlorzoxazone as a metabolic probe. Chlorzoxazone 6-hydroxylation activity was significantly correlated with 4-nitrophenol 2-hydroxylase activity and immunodetected CYP2E1 in 14 human liver samples (r = 0.92 and 0.81, P < 0.001, respectively). Conversely, this catalytic activity was not correlated with CYP 3A or CYP1A activities. Diethyldithiocarbamate (DEDTC), a specific CYP2E1 inhibitor, reduced chlorzoxazone 6-hydroxylase activity by 92.3 +/- 7.6% (n = 14 samples) while ketoconazole, a specific CYP3A inhibitor, reduced this activity by 8.6 +/- 6.3% (n = 14). The residual activity following preincubation with DEDTC was significantly correlated with nifedipine oxidation and tamoxifen N-demethylations, both specific to CYP3A (r = 0.76 and 0.68, respectively). Genetically produced pure human CYP2E1 and 3A4 hydroxylated chlorzoxazone with turnover numbers of 19.7 and 0.14 min(-1), respectively. Furthermore, cytochrome b5 stimulated chlorzoxazone 6-hydroxylation. From examination of the relative liver content of CYP2E1 and 3A, it can be asserted that CYP2E1 is the major enzyme involved in chlorzoxazone 6-hydroxylation and that the contribution of CYP3A is very minor. CYP2E1 activity was evaluated by the plasmatic metabolic ratio 6-OH-CHZ/CHZ (CHZ-MR) measured 2 h after ingestion of 500 mg CHZ. Smoker status did not influence the rate of CHZ hydroxylation. The CHZ-MR was 0.30 +/- 0.13 (mean +/- SD) n = 39 non-smokers versus 0.32 +/- 0.15, n = 75 smokers. This result suggests that CYP1A, inducible by cigarette smoking, is not significantly involved in chlorzoxazone hydroxylation. Women exhibited a slightly lower CHZ-MR than men (0.29 +/- 0.15, n = 44 versus 0.34 +/- 0.15 n = 49, respectively). Obesity increased CHZ-MR, especially in non-insulin-dependent diabetic individuals (0.45 +/- 0.21, n = 13 versus 0.30 +/- 0.15, n = 42 control individuals, P = 0.007). Furthermore, exposure of workers to volatile organics in a shoe factory decreased CHZ-MR (0.19 +/- 0.09, n = 10 Mexican workers versus 0.34 +/- 0.12, n = 16 Mexican control individuals, P = 0.001). Concomitant administration of grapefruit juice (known to be an inhibitor of CYP3A4) with chlorzoxazone did not significantly modify the CHZ metabolic ratio: 0.29 +/- 0.1 versus 0.31 +/- 0.1, for nine control individuals without and with grapefruit juice, respectively. In conclusion, all these results demonstrate that chlorzoxazone is a very selective probe for phenotyping CYP2E1 in humans.     

Differential protein binding of indinavir and saquinavir in matched maternal and umbilical cord plasma

AIMS: To determine whether lower umbilical cord than maternal binding of indinavir and saquinavir contributed to the low cord : maternal (C : M) total concentration ratios reported previously. METHODS: Indinavir and saquinavir unbound fraction (fu) was determined using equilibrium dialysis. Buffer solutions of human serum albumin (HSA) (20.0, 30.0, 40.0 g l(-1)) and alpha(1)-acid glycoprotein (AAG) (0.20, 0.60, 2.00 g l(-1)) were spiked with indinavir (1.00 and 8.00 mg l(-1)) or saquinavir (0.15 and 1.50 mg l(-1)). Matched maternal and umbilical cord plasma was spiked with 1.00 mg l(-1) indinavir (n = 12) or 0.15 mg l(-1) saquinavir (n = 20). Spiked protein/plasma solutions were dialyzed against isotonic phosphate buffer, at 37 degrees C. At equilibrium, indinavir and saquinavir concentrations were quantified, and the f(u) determined. RESULTS: Indinavir and saquinavir demonstrated protein concentration-dependent binding in buffer solutions of HSA and AAG. Indinavir f(u) was significantly higher in umbilical cord (0.53 +/- 0.12) compared with maternal (0.36 +/- 0.11) plasma (95% CI of the difference -0.26, -0.097). Similarly, saquinavir fu was different between umbilical cord (0.0090 +/- 0.0046) and maternal plasma (0.0066 +/- 0.0039) (95% CI of the difference -0.0032, -0.0016). The transplacental AAG concentration gradient contributed significantly to the binding differential of both drugs. CONCLUSIONS: The differential plasma binding of both drugs, which was largely the result of the transplacental AAG concentration gradient, would contribute to the low C : M total plasma concentration ratios observed previously. Unbound concentrations of indinavir and saquinavir are likely to be substantially lower in umbilical cord than maternal plasma.     

Effect of glucagon-like peptide-1(7-36) and exendin-4 on the vascular reactivity in streptozotocin/nicotinamide-induced diabetic rats

We investigated the vascular effects of glucagon-like peptide-1 (GLP-1) and Exendin-4 in type 2 diabetic rat aortae. Studies were performed in a normal control group (NC) (0.2 ml i.p. saline, n = 10), streptozotocin (STZ)/nicotinamide diabetic control group (DC) (a single dose of 80 mg/kg STZ i.p. injection 15 min after administration of 230 mg/kg nicotinamide i.p.), GLP-1 (GLPC) control group (1 microg/kg twice daily i.p. for 1 month, n = 10), Exendin-4 control group (EXC) (0.1 microg/kg twice daily i.p. for 1 month, n = 10), GLP-1-treated diabetic group (GLPT) (1 microg/kg twice daily i.p. for 1 month, n = 10), and Exendin-4-treated diabetic group (EXT) (0.1 microg/kg twice daily i.p. for 1 month, n = 10). One month of GLP-1 and Exendin-4 treatment significantly decreased the blood glucose levels of diabetic rats (113 +/- 2 mg/dl, p < 0.001, and 117 +/- 1 mg/dl, p < 0.001, respectively versus 181 +/- 9 mg/dl in the DC group). Sensitivity (pD2) and maximum response (% Max. Relax) of acetylcholine-stimulated relaxations in the DC group (pD2: 6.73 +/- 0.12 and 55 +/- 6, respectively) were decreased compared with the non-diabetic NC group (pD2: 7.41 +/- 0.25, p < 0.05, and 87 +/- 4, p < 0.01). Treating diabetic rats with GLP-1, pD2 values and with Exendin-4, Max. Relax %values of aortic strips to acetylcholine returned to near non-diabetic NC values (pD2: 7.47 +/- 0.15, p < 0.05, and 87 +/- 3, p < 0.01, respectively). Maximal contractile responses (Emax) to noradrenaline in aortic strips from the diabetic DC group (341 +/- 27 mg tension/mg wet weight) were significantly decreased compared with the non-diabetic NC (540 +/- 66 mg tension/mg wet weight, p < 0.001) and the GLPT group (490 +/- 25 mg tension/mg wet weight, p < 0.05). There were no significant differences in pD2 values of aortic strips to noradrenaline from all groups. Emax to KCl in aortic strips from the DC group (247 +/- 10 mg tension/mg wet weight, p < 0.01) was significantly decreased compared with non-diabetic NC group (327 +/- 26 mg tension/mg wet weight). Treating diabetic rats with GLP-1 (GLPT), Emax values of aortic strips to KCl returned to near non-diabetic NC values (271 +/- 12 mg tension/mg wet weight). GLP-1 and (partially) Exendin-4 treatment could improve the increased blood glucose level and normalize the altered vascular tone in type 2 diabetic rats.     

The effects of verapamil on the electrophysiology of the intact immature mammalian heart

The electrophysiologic effects of intravenous verapamil were compared in six intact neonatal puppies ages 3-15 days and in adult dogs. Utilizing standard intracardiac recording and programmed stimulation techniques, sinus and AV nodal function and atrial and ventricular refractory periods were determined following incremental intravenous doses of 0.075 mg/kg, 0.15 mg/kg, and 0.30 mg/kg of verapamil. In the neonate, intravenous verapamil resulted in a significant increase of the sinus cycle length (37 +/- 6%) but no changes occurred in either the percent sinus node recovery time or the corrected sinus node recovery time. The atrial effective refractory period was prolonged by verapamil (32 +/- 12%). Concerning atrioventricular (AV) nodal function, verapamil produced a small but significant prolongation of resting AH interval (50.5 +/- 2.4 ms (control) versus 57.3 +/- 4.7 ms post 0.30 mg/kg) and a dose-related prolongation of the paced cycle length resulting in AV nodal Wenckebach (170 +/- 12.8 ms control, 190 +/- 10.3 ms post 0.075 mg/kg, 215.8 +/- 13.0 ms post 0.15 mg/kg and 246.7 +/- 22.8 ms post 0.30 mg/kg). The effective refractory period (ERP) and functional refractory period (FRP) of the atrioventricular node (AVN) were prolonged in a dose-dependent fashion (ERP-AVN: 75 +/- 19% increase, FRP-AVN: 42 +/- 14% increase). Retrograde conduction, present in all six neonates, was completely abolished in four of six subjects and significantly prolonged in one other puppy by verapamil. No changes in ventricular refractory periods were observed.(ABSTRACT TRUNCATED AT 250 WORDS)     

Lack of acute effect on lidocaine pharmacokinetics from parenteral nutrition

We tested the hypothesis that amino acids in a parenteral nutrition (PN) solution would result in the decreased metabolism of a model compound, lidocaine. One bolus infusion of lidocaine HCl (1 mg/kg) was administered to seven healthy subjects in association with each of three nutrient regimens: (a) a standard PN solution, (b) 10% dextrose water (D10W), and (c) a meal (control) containing similar fluid volume and caloric, protein, and sodium content as the PN solution. Intravenous nutrients were infused consecutively in a random order at 1 L/12 h. Intravenous and control studies were performed 28 days apart. There was no significant difference in the means (+/- SEM) of total body clearance [7.70 +/- 0.70 (PN) versus 6.78 +/- 0.79 (D10W) versus 7.86 +/- 0.93 (control) ml/min kg], half-life [74.0 +/- 12.2 (PN) versus 89.6 +/- 4.35 (D10W) versus 79.2 +/- 7.22 (control) min], volume of distribution [0.82 +/- 0.15 (PN) versus 0.88 +/- 0.13 (D10W) versus 0.78 +/- 0.13 (control) L/kg], and the fraction of unbound lidocaine in the serum [0.34 +/- 0.025 (PN) versus 0.36 +/- 0.019 (D10W) versus 0.33 +/- 0.020 (control)] among the three nutrient regimens. The concentration-time course of the active metabolite, N-ethylglycyl-2,6-xylidide, did not differ significantly regardless of the nutritional regimen used. Our study indicates that amino acids used for PN do not have any acute effect on lidocaine pharmacokinetics in healthy subjects.     

Photoaffinity labeling of the anionic sites in Caco-2 cells mediating saturable transport of hydrophilic cations ranitidine and famotidine

The H(2) antagonists, ranitidine and famotidine, exhibit saturable absorptive transport across Caco-2 cell monolayers and human intestine via a yet unidentified mechanism. A photoreactive derivative of famotidine has been synthesized and evaluated as a photoaffinity probe for the putative transporter protein(s). The probe irreversibly inhibited ranitidine transport across Caco-2 cell monolayers and irreversibly increased the transepithelial electrical resistance (TEER) after UV activation. Photoaffinity labeling was protected by a molar excess of famotidine.     

Effect of human serum albumin on transplacental transfer of glyburide

Glyburide is a second-generation sulfonylurea hypoglycemic drug used for the treatment of select women with pregestational and gestational diabetes mellitus (GDM). In vitro and in vivo investigations demonstrated its very low transplacental transfer to the fetal circulation. However, the factors influencing its low transfer across the human placenta remain unclear. Therefore, the goal of the current investigation was to determine the effect of human serum albumin (HSA) on the transfer and distribution of glyburide across the human placenta. To achieve this goal, the technique of dual perfusion of the placental lobule was utilized. The effect of HSA on the transfer of glyburide was determined at the range of glyburide to HSA molar ratios of 1:2-1:100. The transfer rate of free/unbound glyburide to the fetal circuit was 73+/-10% of the freely diffusible marker compound antipyrine (AP). Data obtained indicates the dependence of glyburide transfer and its retention by the placental tissue on the concentration of HSA.     

Characterization of drug-protein binding process by employing equilibrium sampling through hollow-fiber supported liquid membrane and Bjerrum and Scatchard plots

The technique equilibrium sampling through membrane (ESTM) was extended to measuring the free drug concentration in solutions of drug and protein. Bjerrum and Scatchard plots were employed for characterizing individual drug binding to pure human blood proteins. Four drugs were investigated as a model system: fluvoxamine and ropivacaine which dominantly bind to alpha-acid glycoprotein (AGP), and R,S-ibuprofen and S-ketoprofen which highly bind to human serum albumin (HSA). The level of drug binding to AGP and HSA relied on drug and protein concentrations. Bjerrum and Scatchard plots revealed high affinity constants (K(a)) at low protein concentration. Both Bjerrum and Scatchard plots of fluvoxamine and ropivacaine binding to AGP showed one specific binding site (n(1)=1) with ropivacaine K(a) value close to 5 times higher than the K(a) of fluvoxamine at 22.9muM AGP concentration. Bjerrum plots of ketoprofen and ibuprofen gave total number of binding sites or bound molecules of 6-7, which did not depend on the drug or protein concentration. Scatchard plots of ketoprofen and ibuprofen exhibited two binding sites (n(1) and n(2)) at 0.15muM and 0.75muM HSA concentrations. On one hand, at 0.15muM HSA, ketoprofen and ibuprofen were bound to site I at n(1)=1.2 and n(1)=1.0, respectively. However, at 0.75muM HSA, ketoprofen and ibuprofen were bound to site I at n(1)=1.2 and n(1)=1.9, respectively. On the other hand, site II, at 0.15muM HSA, interacted with ketoprofen and ibuprofen at n(2)=5.6 and 6.7, respectively. However, at 0.75muM HSA, site II interacted with ketoprofen at n(2)=7.4 and ibuprofen at n(2)=6.2. It would be concluded that, upon mixing ketoprofen and ibuprofen in a HSA solution, a ketoprofen-ibuprofen interaction would most likely occur at site II in HSA.     

Allosteric interaction of the neuromuscular blockers vecuronium and pancuronium with recombinant human muscarinic M2 receptors

Neuromuscular blocking drugs produce muscle weakness by interaction with nicotinic-acetylcholine receptors. Cardiovascular side effects have been reported. In this study the neuromuscular blocking drug vecuronium and the controls gallamine and pancuronium slowed the rate of atropine induced [(3)H]N-methylscopolamine dissociation from Chinese hamster ovary cells expressing recombinant human muscarinic M2 receptors K(off) values min(-1); vecuronium (125 nM), atropine 0.45+/-0.07+blocker 0.04+/-0.02; gallamine (21 nM), atropine 0.42+/-0.05+blocker 0.15+/-0.04; pancuronium(21 nM), atropine 0.36+/-0.03+blocker 0.03+/-0.01). These data indicate that vecuronium, gallamine and pancuronium interact with an allosteric site on the muscarinic M2 receptor (located on the heart) and this may explain some of their cardiac side effects.     

Plasma protein binding of the reversible type A MAO inhibitor cimoxatone (MD 780515)

Binding of a new selective reversible type A MAO inhibitor cimoxatone (MD 780515) to plasma proteins was studied in vitro by equilibrium dialysis. Binding to 580 microM human serum albumin (HSA) and to total plasma proteins was 93-96% and independent of cimoxatone concentration (0.15-207 microM). The drug was mainly bound to HSA with two binding sites and a moderate association constant (K = 2.9 X 10(4) M-1). Free fatty acids did not modify cimoxatone binding to HSA. Cimoxatone was also moderately bound to isolated lipoprotein fractions; alpha 1-acid glycoprotein and gamma-globulins did not play an important role in the binding of cimoxatone. MD 770222, the O-demethyl metabolite, appeared to be bound to HSA at the same binding sites as cimoxatone. However, no interaction occurred between the two compounds for 580 microM HSA. L-Tryptophan, bilirubin, the benzodiazepines flunitrazepam and oxazepam, imipramine and aspirin, did not displace cimoxatone from its binding sites. On the other hand, warfarin and phenylbutazone decreased cimoxatone binding to 29 microM HSA but no interaction occurred with 580 microM HSA.     

Factors influencing the protein binding of vancomycin.

Various factors influencing the protein binding of vancomycin were examined using equilibrium dialysis method. Four per cent human serum albumin (HSA) and/or 0.08 per cent alpha-1-acid glycoprotein (AAG), dissolved in isotonic phosphate buffer, were dialyzed against isotonic phosphate buffer of pH 7.4 using Spectrapor 2 membrane. The protein binding of vancomycin to 0.08 per cent AAG was dependent on vancomycin concentrations; the values ranged from 21.1 per cent at the vancomycin concentration of 20 micrograms ml-1 to 5.30 per cent at 2400 micrograms ml-1. However, binding to 4 per cent HSA was relatively constant, 8.79 +/- 2.43 per cent over a vancomycin concentration range of 20-2400 micrograms ml-1. The values to 4 per cent HSA alone and 0.08 per cent AAG alone did not predict the greater binding of vancomycin in the presence of both proteins, especially at higher concentrations of vancomycin; the values to 4 per cent HSA with 0.08 per cent AAG were constant, 26.3 +/- 3.74 per cent, at the vancomycin concentration range of 20-2400 micrograms ml-1. This suggested an interaction between the proteins, which resulted in enhanced binding of vancomycin. The protein binding of vancomycin to 4 per cent HSA with 0.08 per cent AAG was not influenced by the different incubation temperatures (4 degrees, 22 degrees, and 37 degrees), quantities of heparin (up to 40 units ml-1) or AAG (up to 0.16 per cent), or buffers (isotonic phosphate buffer of pH 7.4, phosphate buffer of pH 7.4 and 0.9 per cent NaCl solution) at the vancomycin concentration of 80 micrograms ml-1. Vancomycin was found to be stable in human serum albumin or in isotonic phosphate buffer of pH 7.4.     

Lomefloxacin promotes the interaction between human serum albumin and transferrin: a mechanistic insight into the emergence of antibiotic's side effects

Human serum albumin (HSA) and serum transferrin (TF) are two drug carrier proteins in vivo. In this study it was investigated how lomefloxacin (LMF) binding affected the HSA-TF interaction using different spectroscopic, calorimetric and molecular modeling techniques. Fluorescence, circular dichroism and synchronous fluorescence revealed that LMF could bind to both proteins, resulting in protein conformational changes. Moreover, HSA and TF could interact so that some fluorescence residues were positioned at the interface and were shielded from quenching by LMF. The interaction between HSA and TF was further confirmed by fluorescence resonance energy transfer. Quantitative analyses of the far-UV CD spectra of the HSA-TF interaction in the presence and absence of LMF revealed secondary structural changes in detail. Resonance light-scattering studies demonstrated that the HSA-TF interaction resulted in a new species with a larger size, and that the presence of LMF could further favor this reaction. Isothermal titration calorimetry revealed that electrostatic interaction was dominant in the absence of LMF, whereas van der Waals forces and hydrogen bonding become significant in its presence. On the other hand, it was found that the binding constant of TF bound to HSA was stronger in the presence of LMF. ANS fluorescence further indicated that hydrophobic interactions play a minor part in the HSA-TF system. Molecular modeling studies confirmed the presence of fluorophore residues, hydrogen bonding and electrostatic interactions at the interface of the HSA-TF complex. It also suggested that the binding sites of LMF were not located there. These data indicate that LMF can modify the interaction between HSA and TF as two model proteins present in serum. The relevance to drugs' side effects, pharmacokinetic of drugs and selection of diagnostic biomarker is discussed.     

Comparative bioavailability of two novel coenzyme Q10 preparations in humans

OBJECTIVE: To determine the absorptive properties of 2 novel coenzyme Q10 preparations, a fast-melting tablet and an effervescent tablet, compared with currently available formulations. MATERIALS AND METHODS: In the first trial, the absorptive properties of 4 different coenzyme Q10 preparations (fast-melting, effervescent, soft gelatin, and powder-filled hard shell) were studied in a randomized, single-dose, crossover study. Twenty-four male subjects were given a 60 mg dose of coenzyme Q10 and plasma coenzyme Q10 was measured over the next 12 hours. Pharmacokinetic properties including area under the curve (AUC), maximum plasma concentration (Cmax), time to maximum plasma concentration (Tmax) and elimination half-life (t 1/2) were measured. In a separate single-dose study, the absorptive characteristics of a different coenzyme Q10 soft gel (Q-Gel) were studied in 6 male subjects. RESULTS: Area under the curve (microg/ml x h) for the fast-melting and effervescent formulations, while marginally greater, was not significantly different when compared to the soft gelatin and powder-filled preparations, 5.4 +/- 1.04 (110%) and 5.5 +/- 0.589 (112%) versus 5.0 +/- 0.859 (102%) and 4.9 +/- 0.812 (100%), respectively. Cmax for the 2 novel formulations was also not statistically different from the soft gelatin or powder-filled preparations, 0.87 +/- 0.14 and 0.86 +/- 0.074 versus 0.70 +/- 0.010 and 0.81 +/- 0.159 (microg/ml). Tmax however, was significantly shorter for the fast-melting and effervescent formulations compared with the soft gel and powder-filled forms, 1.3 +/- 0.348 and 2.0 +/- 0.552 versus 3.7 +/- 0.702 and 4.1 +/- 0.993 (h), respectively. The results of the second trial were similar to those of the powder-filled and soft gel formulations from the first study. CONCLUSIONS: The novel fast-melting and effervescent formulations provide a more rapid delivery of CoQ10 to the blood while exhibiting a similar AUC compared with current formulations. The potential clinical significance of this finding should be further evaluated.     

Plasma protein binding of quinine: binding to human serum albumin, alpha 1-acid glycoprotein and plasma from patients with malaria.

The binding of quinine to human serum albumin (HSA), alpha 1-acid glycoprotein (AAG) and plasma obtained from healthy subjects (10 caucasians and 15 Thais) and from Thai patients with falciparum malaria (n = 20) has been investigated. In healthy volunteers, plasma protein binding expressed as the percentage of unbound quinine was 7.9-31.0% (69-92.1% bound). The mean percentage of unbound quinine found with essentially fatty acid-free HSA (40 g L-1) was 65.4 +/- 1.5% (mean +/- s.d.) and was comparable with the value (66.3 +/- 3.8%, mean +/- s.d.) for Fraction V HSA (40 g L-1). This suggests that fatty acids do not influence the plasma protein binding of quinine. Binding of quinine to 0.7 g L-1 AAG was high (mean unbound 61.0 +/- 5.0%), indicating that quinine is bound primarily to AAG and albumin, although other plasma proteins such as lipoproteins may be involved. The mean percentage of unbound quinine was slightly less in caucasians (14.8 +/- 6.7% unbound), compared with healthy Thai subjects (17.0 +/- 6.7% unbound). The higher binding of quinine in caucasian subjects was associated with a higher plasma AAG concentration observed in caucasians. Mean percentage of unbound quinine was significantly lower in Thai patients with malaria (10.9 +/- 4.0%) than in the healthy Thai subjects. The increase in the extent of quinine binding corresponded with the increase in the acute-phase reactant protein, AAG in the patients with malaria. Overall, when the data were combined there was a significant correlation (r = 0.846, P < 0.005) between the binding ratio (bound/unbound) of quinine and the plasma AAG concentration.(ABSTRACT TRUNCATED AT 250 WORDS)     

Lessons learned from the clinical development of oral peptides

The oral delivery of peptides and proteins has been hampered by an array of obstacles. However, several promising novel oral delivery systems have been developed. This paper reviews the most advanced oral formulation technologies, and highlights key lessons and implications from studies undertaken to date with these oral formulations. Special interest is given to oral salmon calcitonin (CT), glucagon-like peptide-1 (GLP-1), insulin, PYY-(3-36), recombinant human parathyroid hormone (rhPTH(1-31)-NH₂) and PTH(1-34), by different technologies. The issues addressed include (i) interaction with water, (ii) interaction with food, (iii) diurnal variation, (iv) inter- and intra-subject variability, (v) correlation between efficacy and exposure and (vi) key deliverables of different technologies. These key lessons may aid research in the development of other oral formulations.     

Airway retention of materials of different solubility following local intrabronchial deposition in dogs.

We used a gamma camera to monitor the retention and clearance of radiolabeled human serum albumin (HSA), a water-soluble material with molecular weight of 66,000 Daltons, and radiolabeled sulfur colloid (SC), an insoluble submicron (0.22 microm) particle, following localized deposition in a 4-5-mm bronchus in each of five dogs. The average retention time of HSA was significantly greater than that of SC both in the deposition site (23.3 +/- 2.3 vs. 18.2 +/- 2.9 min respectively, p < 0.05) and in the clearance pathway (26.5 +/- 2.0 vs. 22.0 +/- 1.4 min, respectively, p < 0.05). The mean percent retention at the deposition site at 60 minutes post-deposition also was significantly greater for HSA than for SC (33.5 +/- 10.1 vs. 11.6 +/- 4.7%, respectively, p < 0.05). The percentage of HSA which had cleared to the level of the cuffed endotracheal tube was significantly less than that of SC (18.0 +/- 6.7 vs. 35.8 +/- 3.5%, respectively, p < 0.05) at 60 min post-deposition. These findings indicate that a low-permeating water-soluble material such as HSA deposited on the surface of an airway remains in contact with the sensitive airway epithelium to a greater extent than does a solid insoluble particle. Based on our results, we speculate that the slower clearance of HSA compared to SC was likely due to diffusion of a greater portion of the HSA into the periciliary sol layer which may be transported less efficiently than the mucus layer during mucociliary clearance. Additionally, some degree of uptake of HSA by bronchial epithelium may have contributed to its increased retention.     

我国成人肝微粒体的制备和几种细胞色素P450单加氧酶的活性

7例人肝标本均取自我国成年男性,其中5例来自非疾病死亡者,1例为肝血管瘤手术切除的正常肝组织,1例取自交通事故死亡者。该7例人肝微粒体细胞色素P450和细胞色素b5含量分别为0.36±0.08和0.23±0.05nmol·mg^-1蛋白,氨基比林和乙基吗啡N-脱甲基酶活性分别为1.07±0.23和1.82±0.31nmol·mg^-1·min^-1,7-乙氧基香豆素O—脱乙基酶、硝苯吡啶氧化酶和(-)-吡喹酮羟化酶活性分别为0.30±0.10,0.43±0.18和0.69±0.43nmol.mg^-1·min^-1。