• Mass Spectrometry Letters
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• 제8권2호
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• pp.34-38
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• 2017

The purpose of this study was to develop a cocktail approach for the measurement of the permeability of marker compounds, caffeine and propranolol (high permeability), ofloxacin (intermediate), atenolol (low), and quinidine (P-glycoprotein substrate), simultaneously. Then we compared the permeability in Caco-2 cells with that in rat intestinal segments. The difference between individual measurement and cocktail approach was less than 20 %, and the permeabilities of these compounds were similar to those previously reported, suggesting that the cocktail transport study and simultaneous drug analysis were successfully developed and validated in this study. Additionally, in the application of this cocktail method, the permeability of five drugs in rat jejunum was similar to that in ileum but different from that in colon, which was measured using the Ussing chamber system. Moreover, permeability in jejunum and ileum was similar to that in Caco-2 cells. In conclusion, the permeability in Caco-2 cells was equivalent to the permeability in rat jejunum and ileum determined with the Ussing system. Therefore, this newly developed cocktail assay and its application to the Ussing system can be a useful tool for robust and rapid screening for site-specific permeability in rat intestine, thus accelerating the prediction of absorption of new chemical entities.

• Archives of Pharmacal Research
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• 제17권2호
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• pp.57-59
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• 1994

The effect of sodium taurodihydrofusidate (STDHF) on drug permeation across nasal mucosa was studied in vitro using Ussing type diffusion chamber. Disodium cromoglicate (DSCG, M.W.5123) and fluorescein isothiocyanate-dextran (FD) of different molecular wieghts (M.W. 4400-71200) were used as model drugs. Pemeation profiles of DSCG and FDs showed a typical pseudo steady-state curve with short lag time. The pemeability coefficient of FD (M.W. 9400) sigmodially increased with increasing STDHF concentration. It also enhanced the DSCG pemeation. Interestingly the enhancement efficacy was independent of molecular weight of penetrants.

• Animal Bioscience
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• 제34권5호
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• pp.880-885
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• 2021

Objective: Omasum is an important site for the absorption of short chain fatty acids. The major route for the transport of acetate is via sodium hydrogen exchanger (NHE). However, a discrepancy in the symmetry of sodium and acetate transport has been previously reported, the mechanism of which is unclear. In this study, we investigated the possible role of carbonic anhydrase (CA) for this asymmetry. Methods: Omasal tissues were isolated from healthy sheep (N = 3) and divided into four groups; pH 7.4 and 6.4 alone and in combination with Ethoxzolamide. Electrophysiological measurements were made using Ussing chamber and the electrical measurements were made using computer controlled voltage clamp apparatus. Effect(s) of CA inhibitor on acetate and sodium transport flux rate of Na22 and 14C-acetate was measured in three different flux time periods. Data were presented as mean±standard deviation and level of significance was ascertained at p≤0.05. Results: Mucosal to serosal flux of Na (JmsNa) was greater than mucosal to serosal flux of acetate (JmsAc) when the pH was decreased from 7.4 to 6.4. However, the addition of CA inhibitor almost completely abolished this discrepancy (JmsNa ≈ JmsAc). Conclusion: The results of the present study suggest that the additional protons required to drive the NHE were provided by the CA enzyme in the isolated omasal epithelium. The findings of this study also suggest that the functions of CA may be exploited for better absorption in omasum.

• Journal of Photoscience
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• 제3권3호
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• pp.127-132
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• 1996

Calcium has a variety of functions in neuron and muscle cells and blood clotting, especially in the visual system where dark adapted rods cotransport with Na$^+$ into the cell. An influx of Ca$^{2+}$ flows out of the cell through the Na$^+$ - Ca$^{2+}$ exchanger. By using a modified Ussing chamber in order to bring in vivo environment close, we have concluded that Ca$^{2+}$ blocks the activity of guanylate cyclase; in consequence, having an effect on the amplitude of electroretinogram (ERG). We suggest that Ca$^{2+}$ moves between the photoreceptor and the vitreous humor by way of certain Ca$^{2+}$ transport mechanisms. Also, the effect of Zn$^{2+}$ in Ca$^{2+}$ - free ringer solution caused an elevation of amplitude in ERG and a reduction of threshold.

• Archives of Pharmacal Research
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• 제29권7호
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• pp.605-616
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• 2006

A wide variety of drugs and endogenous bioactive amines are organic cations (OCs). Approximately 40% of all conventional drugs on the market are OCs. Thus, the transport of xenobiotics or endogenous OCs in the body has been a subject of considerable interest, since the discovery and cloning of a family of OC transporters, referred to as organic cation transporter (OCTs), and a new subfamily of OCTs, OCTNs, leading to the functional characterization of these transporters in various systems including oocytes and some cell lines. Organic cation transporters are critical in drug absorption, targeting, and disposition of a drug. In this review, the recent advances in the characterization of organic cation transporters and their distribution in the small intestine are discussed. The results of the in vitro transport studies of various OCs in the small intestine using techniques such as isolated brush-border membrane vesicles, Ussing chamber systems and Caco-2 cells are discussed, and in vivo knock-out animal studies are summarized. Such information is essential for predicting pharmacokinetics and pharmacodynamics and in the design and development of new cationic drugs. An understanding of the mechanisms that control the intestinal transport of OCs will clearly aid achieving desirable clinical outcomes.

• Journal of Pharmaceutical Investigation
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• 제24권3호spc1호
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• pp.49-57
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• 1994

The objective of this study was to determine whether Pz-peptide, an enhancer of hydrophilic solute permeability in the intestine, could elevate the paracellular permeability of the cornea and conjunctiva in the pigmented rabbit. The in vitro penetration of four hydrophilic solutes, mannitol (MW 182), fluorescein (MW 376), FD-4 (FITC-dextran, 4 KDa), and FD-10 (FITC-dextran, 10 KDa) across the pigmented rabbit cornea and conjunctiva was studied either in the presence or absence of 3 mM enhancers. Drug penetration was evaluated using the modified Ussing chamber. The conjunctiva was more permeable than the cornea to all four markers. EDTA and cytochalasin B showed higher effects on marker transport than Pz-peptide, but Pz-peptide elevated the corneal transport of mannitol, fluoresein, and FD-4 by 50%, 26%, and 50%, respectively, without affecting FD-10 transport. Possibly due to the leakier nature of the conjunctiva, 3 mM Pz-peptide elevated the transport of only FD-4 by about 45%, without affecting the transport of other markers. Furthermore, the transport of Pz-peptide itself across the cornea and conjunctiva increased with increasing concentration in the 1-5 mM range, suggesting that Pz-peptide enhanced its own permeability, possibly by elevating paracellular permeability. Effects of ion transport inhibitors on Pz-peptide transport were then investigated. PZ-peptide penetration was not changed by mucosal addition of $10\;{\mu}M$ amiloride or $10\;{\mu}M$ hexamethylene amiloride, inhibiting serosal $Na^{+}$ exit by $100\;{\mu}M$ ouabain, or replacing $Na^{+}$ with choline chloride in the mucosal side buffer. These results seggested that Pz-peptide enhanced the paracellular permeability of rabbit cornea and conjunctiva and further indicate that ion transporters were not involved in the Pz-peptide induced elevation of paracellular marker permeability.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1998년도 Proceedings of UNESCO-internetwork Cooperative Regional Seminar and Workshop on Bioassay Guided Isolation of Bioactive Substances from Natural Products and Microbial Products
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• pp.137-137
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• 1998

Stability, efficacy, absorption and toxicity of a new nasal spray formulation including salmon calcitonin were studied in the laboratory animals. After the effects of many excipients on the stability of salmon calcitonin were evaluated using HPLC system, we selected taurine. Our experimental composition of salmon calcitonin contains taurine as a stabilizer and HPMC (hydroxypropylmethyl cellulose) as an adhesive polymer. After intranasal administration of salmon calcitonin formulations, Mia$\^$(R)/, Men$\^$(R)/ and experimental composition, 22 IU to rats, the reduction percentages of calcium concentration in plasma (ΔD%) were 16.3%, 12.9% and 20.8%, respectively. After intranasal administration of Mia$\^$(R)/, Men$\^$(R)/ and experimental composition to rats, C$\sub$MAX/ (205${\pm}$161, 244${\pm}$117, and 330${\pm}$202 pg/$m\ell$, respectively) and AUC (41585${\pm}$22070, 41191${\pm}$19125, and 63357${\pm}$43126 pg. min/$m\ell$, respectively) were calculated. The permeation coefficients 10$\^$-7/,cm/sec) of salmon calcitonin in Mia$\^$(R)/, Men$\^$(R)/ and experimental composition using Ussing chamber with rabbit nasal mucosa were 4.7${\pm}$1.5, 0.75${\pm}$0.4 and 5.3${\pm}$1.1, respectively. The experimental composition with taurine and HPMC was proved to be excellent because it improved the stability of salmon calcitonin and inhanced the absorption of salmon calcitonin and was not irritative to the nasal mucosa.

• Journal of Pharmaceutical Investigation
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• 제26권1호
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• pp.43-53
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• 1996

The objective of this study was to determine whether 4-phenylazobezyloxycarbonyl-Pro-Leu-Gly-Pro-D-Arg (Pz-peptide), an enhancer of hydrophilic solute permeability in the intestine, could elevate the paracellular permeability of hydrophilic drugs across cornea and conjunctiva in the rabbit. The in-vitro penetration of hydrophilic drugs (mannitol, atenolol) and lipophilic drug (propranolol) across the rabbit cornea and conjunctiva was studied either in the presence or absence of 3 mM Pz-peptide. Drug penetration was evaluated using the modified Ussing chamber. The conjunctiva was more permeable than the cornea to all drugs. Pz-peptide showed enhanced effects on the drug transport across cornea and conjunctiva in a concentration dependent manner. Effects or ion transport inhibitor on the mannitol penetration were then investigated. Mannitol penetration was not changed by serosal addition of $100\;{\mu}M$ ouabain, suggesting that $Na^+/K^+$ ion tranporter was not involved in the Pz-peptide induced elevation of paracellular drug permeability. Furthermore, effects of Pz-peptide and EDTA on the transport of atenolol and propranolol into the ocular tissues or blood circulation after its administration into both eyes were investigated. EDTA showed enhanced effect on propranolol transport into the ocular tissues, but Pz-peptide did not show significant difference. Systemic absorption of propranolol by the addition of EDTA or Pz-peptide was not changed. On the other hand, EDTA and Pz-peptide elavated the atenolol transport into the ocular tissues. The transport of atenolol into the blood circulation was also enhanced by the addition of EDTA, but no effect was observed by the addition of Pz-peptide. The above findings suggest that Pz-peptide would be used as an paracellular pathway enahncer of hydrophilic drugs into the eye, without affecting the systemic absortion of topically applied opthalmic drugs.

• Tuberculosis and Respiratory Diseases
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• 제40권3호
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• pp.236-249
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• 1993

연구배경 : 정상 폐상피세포에서는 항상 생성되고 있는 활성산소(oxygen radical)에 의한 유해작용에 노출되어 있고, 이들 유해 산소들은 폐기종과 같은 폐질환의 원인 기전으로 생각되고 있다. 본 연구에서는 이 방법을 이용하여 만든 폐상피세포 단일막에서 전기생리학적인 관점에서 물질의 이동지표인 short-circuit current(Isc)와 조직저항(R)에 대한 활성산소의 하나인 $H_2O_2$(hydrogen peroxide)가 어떤 영향을 미치는지를 연구함으로서 세포생리학적 기전을 구명하고자 한다. 방법 : Tissue culture-treated polycarbonant membrane filter 에서 배양시킨 쥐 제 2 형 폐상피세포 배양 단일막에서 $H_2O_2$의 능동적 이온 이동 (Isc) 과 수동적 용질이동에 대한 조직저항(R)에 미치는 효과를 관찰하였다. 배양 제 3 일과 제 4 일째 단일막을 수정된 Ussing chamber에 설치하고 막 양측에 HEPES-buffered Ringer 용액으로 incubation 하였다. 외부에서 0~100 mM 농도의 $H_2O_2$를 apical 또는 basolateral side에 작용시켜 Isc와 R의 변화를 관찰하였다. 폐상피세포 장벽이 외부의 $H_2O_2$에 대하여 방어작용을 가지는 세포내 catalase 활성도를 측정하고, catalase 억제제인 aminotriazol(ATAZ) 20 mM의 효과도 함께 관찰하였다. 결과 : 이 단일막은 형태학적으로 보아서 in vivo 에서의 포유류 제 1 형 폐상피세포 장벽의 특성을 나타내고 세포들 사이는 tight junction을 이루며(조직저항 R: 2,000 ohm-$cm^2$ 이상) sodium ion의 능동적 이동 (Isc: 5 ${\mu}A/cm^2$)을 보였다. $H_2O_2$는 dose-dependent 양식으로 Isc와 R 모두 감소시켰다. Apical side에 작용하는 $H_2O_2$에 있어서는 60분에 50% 억제하는 농도인 $ED_{50}$는 Isc와 R은 약 4mM 이었으나 basolateral side의 경우는 약 0.04mM 로서 그 작용 강도는 apical에 비하여 약 100배 정도 더 컸다. ATAZ 존재시 apical side의 $ED_{50}$는 0.4mM로 감소하였으나 basolateral side의 경우 변화가 없었다. $H_2O_2$의 제거율은 apical 또는 basolateral side 어느 쪽에 존재하든 같았으며, 세포내 catalase 활성도는세포배양기간이 길어짐에 따라 증가함을 보였다. 결론 : 이상의 실험결과는 basolateral side에 작용하는 $H_2O_2$는 세포내 막구성성분 중 basolateral 측에 존재하는 곳에(예, $Na^+,\;K^+$-APTase) 직접 장애를 미칠 것으로 생각된다. 한편 apical side에 작용하는 $H_2O_2$는 막성분에 도달하기 전에 세포내에 존재하는 catalase에 의하여 대부분 그 작용을 잃게 된다. 결론적으로 Isc와 R로 측정된 폐상피세포 장벽의 특성은 $H_2O_2$에 의하여 손상을 받고 apical side 보다는 basolateral side 측정이 더 손상을 잘 받게 된다.

• The Korean Journal of Physiology and Pharmacology
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• 제24권4호
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• pp.329-338
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• 2020

Rhinorrhea in allergic rhinitis (AR) is characterized by the secretion of electrolytes in the nasal discharge. The secretion of Cl^- and HCO₃^- is mainly regulated by cystic fibrosis transmembrane conductance regulator (CFTR) or via the calcium-activated Cl^- channel anoctamin-1 (ANO1) in nasal gland serous cells. Interleukin-4 (IL-4), which is crucial in the development of allergic inflammation, increases the expression and activity of ANO1 by stimulating histamine receptors. In this study, we investigated ANO1 as a potential therapeutic target for rhinorrhea in AR using an ANO1 inhibitor derived from a natural herb. Ethanolic extracts (30%) of Spirodela polyrhiza (SP_EtOH) and its five major flavonoids constituents were prepared. To elucidate whether the activity of human ANO1 (hANO1) was modulated by SP_EtOH and its chemical constituents, a patch clamp experiment was performed in hANO1-HEK293T cells. Luteolin, one of the major chemical constituents in SP_EtOH, significantly inhibited hANO1 activity in hANO1-HEK293T cells. Further, SP_EtOH and luteolin specifically inhibited the calcium-activated chloride current, but not CFTR current in human airway epithelial Calu-3 cells. Calu-3 cells were cultured to confluency on transwell inserts in the presence of IL-4 to measure the electrolyte transport by Ussing chamber. Luteolin also significantly inhibited the ATP-induced increase in electrolyte transport, which was increased in IL-4 sensitized Calu-3 cells. Our findings indicate that SP_EtOH and luteolin may be suitable candidates for the prevention and treatment of allergic rhinitis. SP_EtOH^- and luteolin-mediated ANO1 regulation provides a basis for the development of novel approaches for the treatment of allergic rhinitis-induced rhinorrhea.