Validation of an In vitro-in vivo Assay System for Evaluation of Transdermal Delivery of Caffeine

2019 ◽  
Vol 9 (1) ◽  
pp. 15-20 ◽  
Author(s):  
Fanni Farner ◽  
Luca Bors ◽  
Ágnes Bajza ◽  
Gellért Karvaly ◽  
István Antal ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Assay System ◽  
Diffusion Cell ◽  
Rat Skin ◽  
Topical Formulation ◽  
Franz Diffusion Cell ◽  
Topical Drugs ◽  
In Vivo Assay

Introduction: Degree of skin penetration of topical drugs and cosmetics is a crucial point concerning their effects and tolerability. For testing drug delivery across the dermal barrier different in vitro and in vivo assays have been developed. Caffeine has been shown to have beneficial effects against skin aging, sunburn and hair-loss, and it is protective against melanoma and non-melanoma type skin cancers. Aim of our study was to set up an assay system to evaluate caffeine penetration from topical formulation into the skin. </P><P> Methods: Franz diffusion cells consisting of either a filter paper or an artificial membrane or rat skin were used as in vitro/ex vivo test systems and transdermal microdialysis in anaesthetized rats was performed as an in vivo assay. </P><P> Results: Results indicate that Franz diffusion cell studies provide a good approximation of the release of caffeine from the formulation but are not able to differentiate between 2% and 4% cream concentrations. The maximum concentrations (Cmax) in case of the 2% cream formulation were 708.3 (2.7 μm pore), 78.7 (0.8 &#181;m pore), 45.3 (0.45 &#181;m pore) and 44.9 (rat skin) &#181;g/7.5 mL, respectively. The in vivo microdialysis experiments were in accordance with the in vitro and ex vivo results and gave more information on the dynamics and follicular and transcellular phases of drug penetration through the layers of the skin. </P><P> Discussion and Conclusion: Taken together, Franz diffusion cell and transdermal microdialysis are a good combination to evaluate caffeine release and penetration into the skin from the formulations tested. This system might also be used for rapid testing of other hydrophilic topical drugs and has a benefit in the prediction for human skin absorption and tolerability studies, in an early phase of drug development.

A Comparative Ex Vivo Study of Plasminogen Activators and Proteases for Fibrinolytic Activity and Side Effects in Rabbits

1977 ◽  
Vol 37 (01) ◽  
pp. 154-161 ◽  
Author(s):  
B. A Janik ◽  
S. E Papaioannou
Keyword(s):  
Side Effects ◽  
Effective Dose ◽  
Fibrinolytic Activity ◽  
Ex Vivo ◽  
Plasminogen Activators ◽  
Assay System ◽  
Coagulation Parameters ◽  
Ex Vivo Assay

SummaryUrokinase, streptokinase, Brinase, trypsin, and SN 687, a bacterial exoprotease, have been evaluated in an ex vivo assay system. These enzymes were injected into rabbits and the fibrinolytic activity as well as other coagulation parameters were measured by in vitro techniques. Dose-response correlations have been made using the euglobulin lysis time as a measure of fibrinolytic activity and the 50% effective dose has been determined for each enzyme. Loading doses, equal to four times the 50% effective dose, were administered to monitor potential toxicity revealing that Brinase, trypsin, and SN 687 were very toxic at this concentration.Having established the 50% effective dose for each enzyme, further testing was conducted where relevant fibrinolytic and coagulation parameters were measured for up to two days following a 50% effective dose bolus injection of each enzyme. Our results have demonstrated that urokinase and streptokinase are plasminogen activators specifically activating the rabbit fibrinolytic system while Brinase, trypsin and SN 687 increase the general proteolytic activity in vivo.The advantages of this ex vivo assay system for evaluating relative fibrinolytic potencies and side effects for plasminogen activators and fibrinolytic proteases have been discussed.

scholarly journals Fucoxanthin-Containing Cream Prevents Epidermal Hyperplasia and UVB-Induced Skin Erythema in Mice

Marine Drugs ◽  
2018 ◽  
Vol 16 (10) ◽  
pp. 378 ◽  
Author(s):  
Azahara Rodríguez-Luna ◽  
Javier Ávila-Román ◽  
María González-Rodríguez ◽  
María Cózar ◽  
Antonio Rabasco ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Antioxidant Properties ◽  
Epidermal Hyperplasia ◽  
Topical Formulation ◽  
Tnf Α ◽  
Skin Erythema ◽  
Cox 2 ◽  
Anti Inflammatory

Microalgae represent a source of bio-active compounds such as carotenoids with potent anti-inflammatory and antioxidant properties. We aimed to investigate the effects of fucoxanthin (FX) in both in vitro and in vivo skin models. Firstly, its anti-inflammatory activity was evaluated in LPS-stimulated THP-1 macrophages and TNF-α-stimulated HaCaT keratinocytes, and its antioxidant activity in UVB-irradiated HaCaT cells. Next, in vitro and ex vivo permeation studies were developed to determine the most suitable formulation for in vivo FX topical application. Then, we evaluated the effects of a FX-containing cream on TPA-induced epidermal hyperplasia in mice, as well as on UVB-induced acute erythema in hairless mice. Our results confirmed the in vitro reduction of TNF-α, IL-6, ROS and LDH production. Since the permeation results showed that cream was the most favourable vehicle, FX-cream was elaborated. This formulation effectively ameliorated TPA-induced hyperplasia, by reducing skin edema, epidermal thickness, MPO activity and COX-2 expression. Moreover, FX-cream reduced UVB-induced erythema through down-regulation of COX-2 and iNOS as well as up-regulation of HO-1 protein via Nrf-2 pathway. In conclusion, FX, administered in a topical formulation, could be a novel natural adjuvant for preventing exacerbations associated with skin inflammatory pathologies as well as protecting skin against UV radiation.

scholarly journals In Vivo Assay of Protein-Protein Interactions in Hin-Mediated DNA Inversion

1998 ◽  
Vol 180 (22) ◽  
pp. 5954-5960 ◽  
Author(s):  
Sun Young Lee ◽  
Hee Jung Lee ◽  
Heejin Lee ◽  
Shukho Kim ◽  
Eun Hee Cho ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Transcriptional Repression ◽  
Assay System ◽  
Host Cells ◽  
Stable Complex ◽  
Protein Protein Interactions ◽  
Dna Inversion ◽  
In Vivo Assay

ABSTRACT In order to form the catalytic nucleoprotein complex called the invertasome in the Hin-mediated DNA inversion reaction, interactions of the DNA-binding proteins Hin and Fis are required. Assays for these protein-protein interactions have been exploited with protein cross-linkers in vitro. In this study, an in vivo assay system that probes protein-protein interactions was developed. The formation of a DNA loop generated by protein interactions resulted in transcriptional repression of an artificially designed operon, which in turn increased the chance of survival of Escherichia colihost cells in a streptomycin-containing medium. Using this system, we were able to assay the Hin-Hin interaction that results in the pairing of the two recombination sites and protein interactions that result in the formation of the invertasome. This assay system also led us to find that an individual Hin dimer bound on a recombination site can form a stable complex with Fis bound on the recombinational enhancer; this finding has never been observed in in vitro studies. Possible pathways toward the formation of the invertasome are discussed based on the assay results for a previously reported Hin mutant.

scholarly journals In Vitro, Ex Vivo, Penetration (EpiDermTM) and in Vivo Dermatokinetics of Ketoconazole Loaded Solid Lipid Nanoparticles for Topical Delivery

2021 ◽  
Author(s):  
Mohhammad Ramzan ◽  
Samuel Gourion-Arsiquaud ◽  
Afzal Hussain ◽  
Jaspreet Singh Gulati ◽  
Qihong Zhang ◽  
...  
Keyword(s):  
Solid Lipid Nanoparticles ◽  
Ex Vivo ◽  
Lipid Nanoparticles ◽  
Laser Scanning Microscopy ◽  
Pharmacokinetic Parameters ◽  
Rat Skin ◽  
Solid Lipid ◽  
Ex Vivo Permeation

Abstract The study focused to optimize, evaluate and investigate mechanistic perspective of ketoconazole (KTZ) loaded solid lipid nanoparticles (KTZ-SLNs) for enhanced permeation across rat skin. KTZ-SLNs were evaluated for size, distribution, zeta potential (ZP), percent entrapment efficiency (%EE), drug release, morphology, thermal behavior (DSC), compatibility (FTIR) and solid state characterization (X-ray diffraction, XRD). Moreover, ex-vivo permeation and drug deposition into rat skin were conducted using Franz diffusion cell. Mechanistic evaluations were confirmed using confocal laser scanning microscopy and vibrational ATR methods using EpiDermTM model. An in vivo dermatokinetics study was performed to ensure KTZ access to the dermal region. Accelerated and photostability studies were conducted at different temperatures (0, 30, and 40 °C) for 12 months. The spherical optimized KOF1 showed optimal particle size (291 nm), and high negative ZP (-27.7 mV). Results of FTIR, DSC, and XRD confirmed compatibility of KTZ with excipients, purity of KTZ & dissolved KTZ in lipid matrix, and amorphous nature of KTZ-SLNs. In-vitro release was found to be slow and sustained whereas ex vivo permeation parameters were significantly high in KTZ-SLNs as compared to drug suspension and marketed product. Drug retention was 10- and -5 fold higher than KTZ-SUS and marketed product, respectively. Pharmacokinetic parameters were improved by SLNs formulation. Confocal raman spectroscopy experiment showed that KTZ-SLNs could penetrate beyond the human stratum corneum into viable epidermis. Fluorescent microscopy confirmed improved penetration of KTZ-SLNs was through human follicular pathway. KTZ-SLNs stable over 12 months under set conditions.

scholarly journals Medicsen Smartpatch: A New Approach to Diabetes Management

2021 ◽  
Vol 5 (Supplement_1) ◽  
pp. A322-A322
Author(s):  
Alejandro Ruiz ◽  
Rafael Danilo García ◽  
Carlos Jorgensen ◽  
Eduardo Jorgensen
Keyword(s):  
Diabetes Management ◽  
In Vivo Model ◽  
Diffusion Cell ◽  
Franz Diffusion Cell ◽  
Glucose Levels ◽  
Inhaled Insulin ◽  
Non Invasive ◽  
Wide Range

Abstract Introduction: Diabetes is a metabolic disorder characterized by a dysregulation of the glucose levels. With insulin being the main drug to be administrated for glucose levels modulation, it needs to be injected subcutaneously with daily injections, which can lead to poor patient compliance, apart from several side-effects. Although other administration methodologies have been investigated (oral or inhaled insulin), they show enough drawbacks to not to be consider as feasible alternatives for diabetes therapy. That′s why Medicsen has developed a Smartpatch that integrates a wide range of technologies, with the purpose of ensuring the correct insulin delivery from the skin′s surface to the bloodstream using a non-invasive and painless drug delivery method through a phenomenon induced by sonophoresis. Materials and Methods: Several in vitro and in vivo tests have been performed to prove the efficacy and safety of the technology, allowing us to collect experimental evidence through different methodologies that demonstrate the therapeutic potential of the device. Among these methodologies, permeability studies using Franz diffusion Cell and swine models (that prove efficacy of the technology) as well as safety studies, for both the insulin and the skin are highlighted. Results: In voltage experiments, the mean time for the disappearance of the membrane potential between the compartments separated by skin was: 334.7 (SD+/-103.6) seconds. Regarding the slope of the voltage line, as an approximation to the transfer speed, an arithmetic mean of (μ)= 0.0164 Mvolts/sec (+/- SD(σ): 0.006) was obtained. No significant differences were found between the circular dichroism spectra of samples (minimum peak at 219nm (sd+/-8.31) and that of the standard, which suggests that the molecular structure of insulin maintains stability. In the same way, HPLC studies shows no variability between the standard and all groups tested. Regardin skin safety, SEM images shows no significant damage to the skin, and ELISA test for TNF α and IL-2, as well as other biochemical tests, show no differences between control and samples. On In vivo experiments with our technology, glucose changes are comparable to those evoked through direct drug injection using conventional syringes. Lastly, the technology proved to be effective in the delivery of insulin through the skin in a non-invasive way, as observed in a Franz Diffusion Cell system and in the in vivo model of blood glucose reduction. Conclusions: Results observed during in vitro and in vivo studies indicate that the technology developed by Medicsen is effective and safe for the patient and the insulin. Following steps, including human trials, will be critical to fully demonstrate its potential in the treatment of diabetes.

The Role of Polyphenols in the Modulation of Plasma Non-Enzymatic Antioxidant Capacity (NEAC)

2012 ◽  
Vol 82 (3) ◽  
pp. 228-232 ◽  
Author(s):  
Mauro Serafini ◽  
Giuseppa Morabito
Keyword(s):  
Antioxidant Capacity ◽  
Dietary Intervention ◽  
Ex Vivo ◽  
The Body ◽  
Dietary Polyphenols ◽  
Enzymatic Antioxidant ◽  
Endogenous Antioxidant

Dietary polyphenols have been shown to scavenge free radicals, modulating cellular redox transcription factors in different in vitro and ex vivo models. Dietary intervention studies have shown that consumption of plant foods modulates plasma Non-Enzymatic Antioxidant Capacity (NEAC), a biomarker of the endogenous antioxidant network, in human subjects. However, the identification of the molecules responsible for this effect are yet to be obtained and evidences of an antioxidant in vivo action of polyphenols are conflicting. There is a clear discrepancy between polyphenols (PP) concentration in body fluids and the extent of increase of plasma NEAC. The low degree of absorption and the extensive metabolism of PP within the body have raised questions about their contribution to the endogenous antioxidant network. This work will discuss the role of polyphenols from galenic preparation, food extracts, and selected dietary sources as modulators of plasma NEAC in humans.

Significance of Platelet β-Adrenoceptors for Platelet Responses In Vivo and In Vitro

1992 ◽  
Vol 68 (06) ◽  
pp. 687-693 ◽  
Author(s):  
P T Larsson ◽  
N H Wallén ◽  
A Martinsson ◽  
N Egberg ◽  
P Hjemdahl
Keyword(s):  
Ex Vivo ◽  
High Dose ◽  
Platelet Aggregability ◽  
Adrenoceptor Agonist ◽  
Dose Infusion ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Factor Antigen

SummaryThe significance of platelet β-adrenoceptors for platelet responses to adrenergic stimuli in vivo and in vitro was studied in healthy volunteers. Low dose infusion of the β-adrenoceptor agonist isoprenaline decreased platelet aggregability in vivo as measured by ex vivo filtragometry. Infusion of adrenaline, a mixed α- and β-adrenoceptor agonist, increased platelet aggregability in vivo markedly, as measured by ex vivo filtragometry and plasma β-thromboglobulin levels. Adrenaline levels were 3–4 nM in venous plasma during infusion. Both adrenaline and high dose isoprenaline elevated plasma von Willebrand factor antigen levels β-Blockade by propranolol did not alter our measures of platelet aggregability at rest or during adrenaline infusions, but inhibited adrenaline-induced increases in vWf:ag. In a model using filtragometry to assess platelet aggregability in whole blood in vitro, propranolol enhanced the proaggregatory actions of 5 nM, but not of 10 nM adrenaline. The present data suggest that β-adrenoceptor stimulation can inhibit platelet function in vivo but that effects of adrenaline at high physiological concentrations are dominated by an α-adrenoceptor mediated proaggregatory action.

In Vivo Effect of Suloctidil as an Antiplatelet Agent

1979 ◽  
Vol 41 (03) ◽  
pp. 465-474 ◽  
Author(s):  
Marcia R Stelzer ◽  
Thomas S Burns ◽  
Robert N Saunders
Keyword(s):  
Ex Vivo ◽  
Antiplatelet Agent ◽  
Ratio Method ◽  
Platelet Aggregate ◽  
Permanent Effect ◽  
Platelet Aggregates ◽  
5 Ht Uptake ◽  
High Doses

SummaryThe relationship between the effects of suloctidil in vivo as an antiplatelet agent and in vitro as a modifier of platelet serotonin (5-HT) parameters was investigated. Suloctidil was found to be effective in reducing platelet aggregates formation in the retired breeder rat as determined using the platelet aggregate ratio method (PAR) with an ED50 of 16.1 mg/kg 24 hours post administration. In contrast to the hypothesis that 5-HT depletion is involved in the anti-aggregatory mechanism of suloctidil, no correlation was found between platelet 5- HT content and this antiplatelet activity. Reduction of platelet 5-HT content required multiple injections of high doses (100 mg/kg/day) of suloctidil. Suloctidil administration for 8 days at 100 mg/kg/day, which lowered platelet 5-HT content by 50%, resulted in no permanent effect on ex vivo platelet 5-HT uptake or thrombin-induced release, nor alteration in the plasma 5-HT level. However, these platelets exhibited a short-lived, significant increase in percent leakage of 5-HT after 30 minutes of incubation. Therefore, suloctidil treatment at high doses may with time result in platelet 5-HT depletion, however this effect is probably not related to the primary anti-aggregatory activity of the drug.

Organische Nitrate und Thrombozytenfunktion

1988 ◽  
Vol 08 (02) ◽  
pp. 90-99 ◽  
Author(s):  
H. Schröder ◽  
K. Schrör
Keyword(s):  
Endothelial Cell ◽  
Angina Pectoris ◽  
Ex Vivo

ZusammenfassungOrganische Nitrate unterschiedlicher chemischer Struktur sowie Nitroprussidnatrium und Molsidomin (bzw. ihre biologisch aktiven Metaboliten) können die (primäre) Aggregation und Sekretion von Humanthrombozyten in vitro und ex vivo hemmen. Eine solche Wirkung wird für Molsidomin (SIN-1) und Nitroprussidnatrium in vitro in Konzentrationen beobachtet, die in der gleichen Größenordnung liegen wie die vasodilatierenden Effekte der Substanzen. Dagegen sind für eine direkte Antiplättchenwirkung organischer Nitrate (Glyzeryltrinitrat, Isosorbiddinitr at, Isosorbidmononitrate, Teopranitol) in vitro Konzentrationen erforderlich, die ca. 100- bis 1000fach höher sind als die Plasmaspiegel der Substanzen nach therapeutischer Dosierung bzw. die Konzentrationen, die isolierte Gefäßstreifen relaxieren. Als gemeinsamer Wirkungsmechanismus der direkten thrombozy-tenfunktionshemmenden und gefäßerweiternden Wirkung all dieser Substanzen kann heute eine Stickoxid-(NO)-vermittelte Stimulation der cGMP-Bildung angenommen werden, das aus organischen Nitraten als »Pro-drug« entsteht. Die Freisetzung von NO, eines »endothelial cell-derived relaxing factors« (EDRF) aus Nitroprussidnatrium und SIN-1 erfolgt spontan. Dagegen erfordert die Freisetzung von NO aus organischen Nitraten einen enzymatischen Stoffwechselweg, der in isolierten Thrombozyten nicht vorhanden ist. Eine Antiplättchenwirkung organischer Nitrate in vivo bzw. ex vivo wird daher über die Stimulation eines endothelialen, thrombozyteninhibitorischen Faktors erklärt. Hierbei sind Prostazyklin sowie ein bisher unbekannter Endothel-zellfaktor neben einer synergistischen Wirkung organischer Nitrate mit endogenem Prostazyklin in Diskussion. Eine thrombozytenfunktionshemmen-de Wirkung organischer Nitrate könnte in Kombination mit ihren hämody-namischen Effekten auch für die an-tianginöse Wirkung in der Klinik bedeutsam sein, insbesondere zur Verhinderung vasospastischer Zustände bei der instabilen Angina pectoris.

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