Simultaneous determination of digoxin and permeability markers in rat in situ intestinal perfusion samples by RP-HPLC

Varma, Manthena V.; Kapoor, Namita; Sarkar, Mahua; Panchagnula, Ramesh

doi:10.1016/j.jchromb.2004.09.047

Cited by 47 publications

(31 citation statements)

References 19 publications

(25 reference statements)

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“…PR is a non-absorbable, non-toxic pH indicator component of many growth media formulations (Lewis, 2002;Rathore et al, 2008), which is also commonly used as a paracellular permeability marker in many epithelial cell studies (Azenha et al, 2004;Paye et al, 2007;Peixoto and Collares-Buzato, 2005). By using high-performance liquid chromatography (HPLC), extremely low concentrations (10ngml -1 ) of the small molecule can be detected (Varma et al, 2004).…”

Section: Ter and Pr Diffusion (Prd) Measurementmentioning

confidence: 99%

Claudin 28b and F-actin are involved in rainbow trout gill pavement cell tight junction remodeling under osmotic stress

Sandbichler

Egg

Schwerte

et al. 2011

Journal of Experimental Biology

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SUMMARYPermeability of rainbow trout gill pavement cells cultured on permeable supports (single seeded inserts) changes upon exposure to freshwater or treatment with cortisol. The molecular components of this change are largely unknown, but tight junctions that regulate the paracellular pathway are prime candidates in this adaptational process. Using differential display polymerase chain reaction we found a set of 17 differentially regulated genes in trout pavement cells that had been exposed to freshwater apically for 24h. Five genes were related to the cell-cell contact. One of these genes was isolated and identified as encoding claudin 28b, an integral component of the tight junction. Immunohistochemical reactivity to claudin 28b protein was concentrated in a circumferential ring colocalized to the cortical F-actin ring. To study the contribution of this isoform to changes in transepithelial resistance and Phenol Red diffusion under apical hypo-or hyperosmotic exposure we quantified the fluorescence signal of this claudin isoform in immunohistochemical stainings together with the fluorescence of phalloidin-probed F-actin. Upon hypoosmotic stress claudin 28b fluorescence and epithelial tightness remained stable. Under hyperosmotic stress, the presence of claudin 28b at the junction significantly decreased, and epithelial tightness was severely reduced. Cortical F-actin fluorescence increased upon hypo-osmotic stress, whereas hyperosmotic stress led to a separation of cortical F-actin rings and the number of apical crypt-like pores increased. Addition of cortisol to the basolateral medium attenuated cortical F-actin separation and pore formation during hyperosmotic stress and reduced claudin 28b in junctions except after recovery of cells from exposure to freshwater. Our results showed that short-term salinity stress response in cultured trout gill cells was dependent on a dynamic remodeling of tight junctions, which involves claudin 28b and the supporting F-actin ring.

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Section: Ter and Pr Diffusion (Prd) Measurementmentioning

confidence: 99%

Claudin 28b and F-actin are involved in rainbow trout gill pavement cell tight junction remodeling under osmotic stress

Sandbichler

Egg

Schwerte

et al. 2011

Journal of Experimental Biology

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“…Intestinal permeability for a variety of drugs obtained from in situ single pass intestinal perfusion experiments have shown excellent correlation with human perfusion studies (11,22). Also, in situ intestinal permeability of nine drugs were well correlated with corresponding human results in terms of whether these drugs being actively absorbed or they are subject to simple passive diffusion (12). In addition, permeability parameters obtained from in situ intestinal perfusion model provided a better prediction of human absorption than the cell-based assays (23).…”

Section: Discussionmentioning

confidence: 81%

“…In situ intestinal perfusion in rats also known as single pass intestinal perfusion is a commonly used technique for assessment of permeability of drugs through intestines while keeping intact blood supply to the intestinal tract throughout the experiment, making it a robust tool for simulating real in vivo conditions following oral drug administration (10)(11)(12)(13). The improved form of this technique, where the whole small intestines is used for more accurate determination of drug's permeability, especially for low permeability compounds, has been recently introduced in our lab and published elsewhere (14).…”

Section: Introductionmentioning

confidence: 99%

In Situ Intestinal Perfusion of Irinotecan: Application to P-gp Mediated Drug Interaction and Introduction of an Improved HPLC Assay

Rabba

Xue

et al. 2011

J Pharm Pharm Sci

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-Purpose:To determine experimentally the intestinal permeability of the anticancer prodrug irinotecan, and to quantify the amount of its cytotoxic metabolite SN-38 that is intestinally excreted (exsorbed) as a predictor of intestinal toxicity, and to assess the effect of p-glycoprotein (p-gp) inhibitors (verapamil as a model) on the permeability and toxicity of irinotecan. Methods: Single pass intestinal perfusion of rat's whole length small intestines is applied to assess the permeability of the parent drug and quantify the intestinally excreted metabolite. The perfusion solution contained 30μg/ml of irinotecan (control group) without or with verapamil (verapamil group). A simple reversed phase HPLC method with UV detection is developed and validated for simultaneous determination of irinotecan and SN-38 using camptothecin as an internal standard. Results: HPLC-UV method found to be simple, specific, accurate, and precise. Effective permeability coefficient of irinotecan found to be 4.9±1.7  10 -3 mm/min and was doubled in verapamil group (P=0.007). Average cumulative amount of SN-38 exsorbed found to be 29 ng/cm over 2 hours perfusion time which was decreased to 15 ng/cm in verapamil group (P=0.016). Conculsions: in situ intestinal perfusion method was successfully applied to quantify the permeability of irinotecan and the exsorption of SN-38 in the same experiment, in a manner that robustly reflects real in vivo situation. P-gp inhibition using verapamil found to significantly enhance the intestinal permeability of irinotecan and potentially decrease the intestinal toxicity due to SN-38 exposure.

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“…Accuracy can be defined as the percent of the recovered analyte from a known added quantity [9]. Data from nine samples representing three concentration levels covering the obtained linearity range (10, 30 and 50 µg/mL for the HPLC method and 10, 20 and 30 µg/mL for the UV-spectrophotometric method) was used to assess accuracy of both methods, so the previously mentioned procedures under linearity were carried out then the concentration of SFV samples was calculated using the corresponding regression equation.…”

Section: Accuracymentioning

confidence: 99%

Simple chromatographic and spectrophotometric determination of sofosbuvir in pure and tablet forms

Abdel-Gawad¹

2016

Eur. J. Chem.

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Two methods, a reversed phase high-performance liquid chromatographic (RP-HPLC) method and a direct ultra-violet spectrophotometric method, were adopted and validated for the quantification of sofosbuvir, which is a new antiviral agent used for treatment of patients with hepatitis C virus (HCV). Validation parameters such as linearity, accuracy, precision, specificity, limits of detection and quantification were determined according to the guidelines of International Conference on Harmonization (ICH)-Q2B. The RP-HPLC method was applied on Hypersil TM ODS C18 column (150×4.6 mm, 5 µm) as a stationary phase. The mobile phase was optimized according to the polarity of the studied drug. It was methanol: acetonitrile (90:10, v:v), pumped using an isocratic mode with flow rate of 1 mL/min and UV detection at 260 nm. The UV spectrophotometric method was performed for the studied drug at 260 nm. The calibration curves were linear in the ranges of 2-60 µg/mL and 5-40 µg/mL for the RP-HPLC and UV spectrophotometric methods, respectively. The proposed methods are accurate, sensitive and precise, so they can be successfully adopted for the reliable determination of sofosbuvir content in its tablet form.

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Simultaneous determination of digoxin and permeability markers in rat in situ intestinal perfusion samples by RP-HPLC

Cited by 47 publications

References 19 publications

Claudin 28b and F-actin are involved in rainbow trout gill pavement cell tight junction remodeling under osmotic stress

Claudin 28b and F-actin are involved in rainbow trout gill pavement cell tight junction remodeling under osmotic stress

In Situ Intestinal Perfusion of Irinotecan: Application to P-gp Mediated Drug Interaction and Introduction of an Improved HPLC Assay

Simple chromatographic and spectrophotometric determination of sofosbuvir in pure and tablet forms

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